Competitive assays for determining the effectiveness of a human immunodeficiency virus type 2 (HIV-2) antiviral agent, employing peptides and proteins of HIV-2

Information

  • Patent Grant
  • 5976785
  • Patent Number
    5,976,785
  • Date Filed
    Friday, December 20, 1991
    32 years ago
  • Date Issued
    Tuesday, November 2, 1999
    24 years ago
Abstract
A method for diagnosing an HIV-2 (LAV-II) infection and a kit containing reagents for the same is disclosed. These reagents include cDNA probes which are capable of hybridizing to at least a portion of the genome of HIV-2. In one embodiment, the DNA probes are capable of hybridizing to the entire genome of HIV-2. These reagents also include polypeptides encoded by some of these DNA sequences.
Description

The invention relates to cloned DNA sequences analogous to the genomic RNA of a virus known as Lymphadenopathy-Associated Virus II ("LAV-II"), a process for the preparation of these cloned DNA sequences, and their use as probes in diagnostic kits. In one embodiment, the invention relates to a cloned DNA sequence analogous to the entire genomic RNA of HIV-2 and its use as a probe. The invention also relates to polypeptides with amino acid sequences encoded by these cloned DNA sequences and the use of these polypeptides in diagnostic kits.
According to recently adopted nomenclature, as reported in Nature, May 1986, a substantially-identical group of retroviruses which has been identified as one causative agent of AIDS are now referred to as Human Immunodeficiency Viruses I (HIV-1). This previously-described group of retroviruses includes Lymphadenopathy-Associated Virus I (LAV-I), Human T-cell Lymphotropic Virus-III (HTLV-III), and AIDS-Related Virus (ARV).
Lymphadenopathy-Associated Virus II has been described in U.S. application Ser. No. 835,228, which was filed Mar. 3, 1986, and is specifically incorporated herein by reference. Because LAV-II is a second, distinct causative agent of AIDS, LAV-II properly is classifiable as a Human Immunodeficiency Virus II (HIV-2). Therefore, "LAV-II" as used hereinafter describes a particular genus of HIV-2 isolates.
While HIV-2 is related to HIV-1 by its morphology, its tropism and its in vitro cytopathic effect on CD4 (T4) positive cell lines and lymphocytes, HIV-2 differs from previously described human retroviruses known to be responsible for AIDS. Moreover, the proteins of HIV-1 and 2 have different sizes and their serological cross-reactivity is restricted mostly to the major core protein, as the envelope glycoproteins of HIV-2 are not immune precipitated by HIV-l-positive sera except in some cases where very faint cross-reactivity can be detected. Since a significant proportion of the HIV infected patients lack antibodies to the major core protein of their infecting virus, it is important to include antigens to both HIV-1 and HIV-2 in an effective serum test for the diagnosis of the infection by these viruses.
HIV-2 was first discovered in the course of serological research on patients native to Guinea-Bissau who exhibited clinical and immunological symptoms of AIDS and from whom sero-negative or weakly sero-positive reactions to tests using an HIV-1 lysate were obtained. Further clinical studies on these patients isolated viruses which were subsequently named "LAV-II."One LAV-II isolate, subsequently referred to as LAV-II MIR, was deposited at the Collection Nationale des Cultures de Micro-Organismes (CNCM) at the Institut Pasteur in Paris, France on Dec. 19, 1985 under Accession No. I-502 and has also been deposited at the British ECA CC under No. 87.001.001 on Jan. 9, 1987. A second LAV-II isolate was deposited at CNCM on Feb. 21, 1986 under Accession No. I-532 and has also been deposited at the British ECA CC under No. 87.001.002 on Jan. 9, 1987. This second isolate has been subsequently referred to as LAV-II ROD. Other isolates deposited at the CNCM on Dec. 19, 1986 are HIV-2 IRMO (No. I-642) and HIV-2 EHO (No. I-643). Several additional isolates have been obtained from West African patients, some of whom have AIDS, others with AIDS-related conditions and others with no AIDS symptoms. All of these viruses have been isolated on normal human lymphocyte cultures and some of them were thereafter propagated on lymphoid tumor cell lines such as CEM and MOLT.
Due to the sero-negative or weak sero-positive results obtained when using kits designed to identify HIV-1 infections in the diagnosis of these new patients with HIV-2 disease, it has been necessary to devise a new diagnostic kit capable of detecting HIV-2 infection, either by itself or in combination with an HIV-1 infection. The present inventors have, through the development of cloned DNA sequences analogous to at least a portion of the genomic RNA of LAV-II ROD viruses, created the materials necessary for the development of such kits.
SUMMARY OF THE INVENTION
As noted previously, the present invention relates to the cloned nucleotide sequences homologous or identical to at least a portion of the genomic RNA of HIV-2 viruses and to polypeptides encoded by the same. The present invention also relates to kits capable of diagnosing an HIV-2 infection.
Thus, a main object of the present invention is to provide a kit capable of diagnosing an infection caused by the HIV-2 virus. This kit may operate by detecting at least a portion of the RNA genome of the HIV-2 virus or the provirus present in the infected cells through hybridization with a DNA probe or it may operate through the immunodiagnostic detection of polypeptides unique to the HIV-2 virus.
Additional objects and advantages of the present invention will be set forth in part in the description which follows, or may be learned from practice of the invention. The objects and advantages may be realized and attained by means of the instrumentalities and combinations particularly pointed out in the appended claims.
To achieve these objects and in accordance with the purposes of the present invention, cloned DNA sequences related to the entire genomic RNA of the LAV-II virus are set forth. These sequences are analogous specifically to the entire genome of the LAV-II ROD strain.
To further achieve the objects and in accordance with the purposes of the present invention, a kit capable of diagnosing an HIV-2 infection is described. This kit, in one embodiment, contains the cloned DNA sequences of this invention which are capable of hybridizing to viral RNA or analogous DNA sequences to indicate the presence of an HIV-2 infection. Different diagnostic techniques can be used which include, but are not limited to: (1) Southern blot procedures to identify viral DNA which may or may not be digested with restriction enzymes; (2) Northern blot techniques to identify viral RNA extracted from cells; and (3) dot blot techniques, i.e., direct filtration of the sample through an ad hoc membrane such as nitrocellulose or nylon without previous separation on agarose gel. Suitable material for dot blot technique could be obtained from body fluids including, but not limited to, serum and plasma, supernatants from culture cells, or cytoplasmic extracts obtained after cell lysis and removal of membranes and nuclei of the cells by ultra-centrifugation as accomplished in the "CYTODOT" procedure as described in a booklet published by Schleicher and Schull.
In an alternate embodiment, the kit contains the polypeptides created using these cloned DNA sequences. These polypeptides are capable of reacting with antibodies to the HIV-2 virus present in sera of infected individuals, thus yielding an immunodiagnostic complex.
In accordance with a further object of the present invention, a peptide is provided as described above, either along or conjugated to a carrier molecule, the peptide being capable of eliciting the production of an antibody to the peptide, and said antibody is capable of forming an effective immunocomplex with the entire HIV-2 retrovirus or with its corresponding proteins.
To further achieve the objects of the invention, a vaccinating agent is provided which comprises at least one peptide selected from the polypeptide expression products of the viral DNA in admixture with suitable carriers, adjuvents stabilizers.
It is understood that both the foregoing general description and the following detailed description are exemplary and explanatory only and are not restrictive of the invention as claimed. The accompanying drawings, which are incorporated in and constitute a part of the specification, illustrate one embodiment of the invention and, together with the description, serve to explain the principles of the invention.





BRIEF DESCRIPTION OF THE DRAWINGS
FIG. 1 generally depicts the nucleotide sequence of a cloned complementary DNA (cDNA) to the genomic RNA of HIV-2. FIG. 1A depicts the genetic organization of HIV-1, position of the HIV-1 HindII fragment used as a probe to screen the cDNA library, and restriction map of the HIV-2 cDNA clone, E2. FIG. 1B depicts the nucleotide sequence of the 3' end of HIV-2. The corresponding region of the HIV-1 LTR was aligned using the Wilbur and Lipman algorithm (window: 10; K-tuple: 7; gap penalty: 3) as described by Wilbur and Lipman in Proc. Natl. Acad. Sci. USA 80: 726-730 (1983), specifically incorporated herein by reference. The U3-R junction in HIV-1 is indicated and the poly A addition signal and potential TATA promoter regions are boxed. In FIG. 1B, the symbols B, H, Ps and Pv refer to the restriction sites BamHI, HindIII, PstI and PvuII, respectively.
FIG. 2 generally depicts the HIV-2 specificity of the E2 clone. FIGS. 2A and B are lines drawing representing Southern blots of DNA extracted from CEM cells infected with the following isolates: HIV-2.sub.ROD (a,c), HIV-2.sub.DUL (b,d), and HIV-1.sub.BRU (e.f). DNA in lanes a,b,f was Pst I digested; in c,d,e DNA was undigested. FIGS. 2C and D are line drawing representing dot blot hybridization of pelleted virions from CEM cells infected by the HIV-1.sub.BRU (1), Simian Immunodeficiency Virus (SIV) isolate Mm 142-83 (3), HIV-2.sub.DUL (4), HIV-2.sub.ROD (5), and HIV-1.sub.ELI (6). Dot 2 is a pellet from an equivalent volume of supernatant from uninfected CEM. Thus, FIGS. 2A and C depict hybridization with the HIV-2 cDNA (E2) and FIGS. 2B and D depict hybridization to an HIV-1 probe consisting of a 9 Kb SacI insert from HIV-1 BRU(clone lambda J 19).
FIG. 3 generally depicts a restriction map of the HIV-2 ROD genome and its homology to HIV-1. FIG. 3A specifically depicts the organization of three recombinant phage lambda clones, ROD 4, ROD 27, and ROD 35. In FIG. 3A, the open boxes represent viral sequences, the LTR are filled, and the dotted boxes represent cellular flanking sequences (not mapped). Only some characteristic restriction enzyme sites are indicated. .lambda.ROD 27 and .lambda.ROD 35 are derived from integrated proviruses while .lambda.ROD 4 is derived from a circular viral DNA. The portion of the lambda clones that hybridzes to the cDNA E2 is indicated below the maps. A restriction map of the .lambda.ROD isolate was reconstructed from these three lambda clones. In this map, the restriction sites are identified as follows: B: BamHI; E: EcoRI; H: HindIII; K: KpnI; Ps: PstI; Pv: PvuII; S: SacI; X: XbaI. R and L are the right and left BamHI arms of the lambda L47.1 vector.
FIG. 3B specifically depicts dots 1-11 which correspond to the single-stranded DNA form of M13 subclones from the HIV-1.sub.BRU cloned genome (.lambda.J19). Their size and position on the HIV-1 genome, determined by sequencing is shown below the figure. Dot 12 is a control containing lambda phage DNA. The dot-blot was hybridized in low stringency conditions as described in Example 1 with the complete lambda .lambda.ROD 4 clone as a probe, and successively washed in 2.times. SSC, 0.1% SDS at 25.degree. C. (Tm -42.degree. C.), 4.times. SSC, 0.1% SDS at 60.degree. C. (Tm -20.degree. C.), and 0.1.times. SSC, 0.1% SDS at 60.degree. C. (Tm -3.degree. C.) and exposed overnight. A duplicate dot blot was hybridized and washed in stringent conditions (as described in Example 2) with the labelled lambda J19 clone carrying the complete HIV-1.sub.BRU genome. HIV-1 and HIV-2 probes were labelled the same specific activity (10.sup.8 cpm/g.).
FIG. 4 generally depicts the restriction map polymorphism in different HIV-2 isolates and shows comparison of HIV-2 to SIV. FIG. 4A is a line drawing depicting DNA (20 .mu.g. per lane) from CEM cells infected by the isolate HIV-2.sub.DUL (panel 1) or peripheral blood lymphocytes (PBL) infected by the isolates HIV-2.sub.GOM (panel 2) and HIV-2.sub.MIR (panel 3) digested with: EcoRI (a), PstI (b), and HindIII (c). Much less viral DNA was obtained with HIV-2 isolates propagated on PBL. Hybridization and washing were in stringent conditions, as described in Example 2, with 10.sup.6 cpm/ml. of each of the E2 insert (cDNA) and the 5 kb. HindIII fragment of .lambda.ROD 4, labelled to 10.sup.9 cpm/.mu.g.
FIG. 4B is a line drawing depicting DNA from HUT 78 (a human T lymphoid cell line) cells infected with STLV3 MAC isolate Mm 142-83. The same amounts of DNA and enzymes were used as indicated in panel A. Hybridization was performed with the same probe as in A, but in non-stringent conditions. As described in Example 1 washing was for one hour in 2.times. SSC, 0.1% SDS at 40.degree. C. (panel 1) and after exposure, the same filter was re-washed in 0.1.times. SSC, 0.1% SDS at 60.degree. C. (panel 2). The autoradiographs were obtained after overnight exposition with intensifying screens.
FIG. 5 depicts the position of derived plasmids from .lambda.ROD 27, .lambda.ROD 35 and .lambda.ROD 4.





DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS
Reference will now be made in detail to the presently preferred embodiments of the invention, which, together with the following examples, serve to explain the principles of the invention.
The genetic structure of the HIV-2 virus has been analyzed by molecular cloning according to the method set forth herein and in the Examples. A restriction map of the genome of this virus is included in FIG. 5. In addition, the partial sequence of a CDNA complementary to the genomic RNA of the virus has been determined. This cDNA sequence information is included in FIG. 1.
Also contained herein is data describing the molecular cloning of the complete 9.5 kb genome of HIV-2, data describing the observation of restriction map polymorphism between different isolates, and an analysis of the relationship between HIV-2 and other human and simian retroviruses. From the totality of these data, diagnostic probes can be discerned and prepared.
Generally, to practice one embodiment of the present invention, a series of filter hybridizations of the HIV-2 RNA genome with probes derived from the complete cloned HIV-1 genome and from the gag and pol genes were conducted. These hybridizations yielded only extremely weak signals even in conditions of very low stringency of hydridization and washing. Thus, it was found to be difficult to assess the amount of HIV-2 viral and proviral DNA in infected cells by Southern blot techniques.
Therefore, a complementary DNA (cDNA) to the HIV-2 genomic RNA initially was cloned in order to provide a specific hybridization probe. To construct this cDNA, an oligo (dT) probed cDNA first-strand was made in a detergent-activated endogenous reaction using HIV-2 reverse transcriptase with virions purified from supernatants of infected CEM cells. The CEM cell line is a lymphoblastoid CD4+ cell line described by G. E. Foley et al. in Cancer 18: 522-529 (1965), specifically incorporated herein by reference. The CEM cells used were infected with the isolate ROD and were continuously producing high amounts of HIV-2.
After second-strand synthesis, the cDNAs were inserted into the M 13 tg 130 bacteriophage vector. A collection of 10.sup.4 M13 recombinant phages was obtained and screened in situ with an HIV-1 probe spanning 1.5 kb. of the 3' end of the LAVBRU isolate (depicted in FIG. 1A). Some 50 positive plaques were detected, purified, and characterized by end sequencing and cross-hybridizing the inserts. This procedure is described in more detail in Example 1 and in FIG. 1.
The different clones were found to be complementary to the 3' end of a polyadenylated RNA having the AATAAA signal about 20 nucleotides upstream of the poly A tail, as found in the long terminal repeat (LTR) of HIV-1. The LTR region of HIV-1 has been described by S. Wain Hobson et al. in Cell 40: 9-17 (1985), specifically incorporated herein by reference. The portion of the HIV-2 LTR that was sequenced was related only distantly to the homologous domain in HIV-1 as demonstrated in FIG. 1B. Indeed, only about 50% of the nucleotides could be aligned and about a hundred insertions/deletions need to be introduced in comparison, the homology of the corresponding domains in HIV-1 isolates from USA and Africa is greater than 95% and no insertions or deletions are seen.
The largest insert of this group of M13 clones was a 2 kb. clone designated E2. Clone E2 was used as a probe to demonstrate its HIV-2 specificity in a series of filter hybridization experiments. Firstly, this probe could detect the genomic RNA of HIV-2 but not HIV-1 in stringent conditions as shown in FIGS. 2, C and D. Secondly, positive signals were detected in Southern blots of DNA from cells infected with the ROD isolate as well as other isolates of HIV-2 as shown in FIG. 2, A and FIG. 4, A. No signal was detected with DNA from uninfected cells or HIV-1 infected cells, confirming the exogenous nature of HIV-2. In undigested DNA from HIV-2 infected cells, an approximately 10 kb. species, probably corresponding to linear unintegrated viral DNA, was principally detected along with a species with an apparent size of 6 kb., likely to be the circular form of the viral DNA. Conversely, rehybridization of the same filter with an HIV-1 probe under stringent conditions showed hybridization to HIV-1 infected cells only as depicted in FIG. 2, B.
To isolate the remainder of the genome of HIV-2, a genomic library in lambda phage L47.1 was constructed. Lambda phage L47.1 has been described by W. A. M. Loenen et al. in Gene 10: 249-259 (1980), specifically incorporated herein by reference. The genomic library was constructed with a partial Sau3AI restriction digest of the DNA from the CEM cell line infected with HIV-2.sub.ROD.
About 2.times.10.sup.6 recombinant plaques were screened in situ with labelled insert from the E2 cDNA clone. Ten recombinant phages were detected and plaque purified. Of these phages, three were characterized by restriction mapping and Southern blot hybridization with the E2 insert and probes from its 3' end (LTR) or 5' end (envelope), as well as with HIV-1 subgenomic probes. In this instance, HIV-1 probes were used under non-stringent conditions.
A clone carrying a 9.5 kb. insert and derived from a circular viral DNA was identified as containing the complete genome and designated .lambda.ROD 4. Two other clones, .lambda.ROD 27 and .lambda.ROD 35 were derived from integrated proviruses and found to carry an LTR and cellular flanking sequences and a portion of the viral coding sequences as shown in FIG. 3, A.
Fragments of the lambda clones were subcloned into a plasmid vector p UC 18.
Plasmid pROD 27-5' is derived from .lambda.ROD 27 and contains the 5' 2 Kb of the HIV-2 genome and cellular flanking sequences (5' LTR and 5' viral coding sequences to the EcoRI site). Plasmid p ROD 4-8 is derived from .lambda.ROD 4 and contains the about 5 Kb HindIII fragment that is the central part of the HIV-2 genome.
Plasmid pROD 27-5' and p ROD 4.8 inserts overlap.
Plasmid pROD 4.7 contains a HindIII 1.8 Kb fragment from .lambda.ROD 4. This fragment is located 3' to the fragment subcloned into pROD 4.8 and contains about 0.8 Kb of viral coding sequences and the part of the lambda phage (.lambda.L47.1) left arm located between the BamHI and HindIII cloning sites.
Plasmid pROD 35 contains all the HIV-2 coding sequences 3' to the EcoRI site, the 3' LTR and about 4 Kb of cellular flanking sequences.
Plasmid pROD 27-5' and pROD 35 in E. coli strain HB 101 are deposited respectively under No. I-626 and I-633 at the CNCM, and have also been deposited at the NCIB (British Collection). These plasmids are depicted in FIG. 5. Plasmids pROD 4-7 and pROD 4-8 in E. coli strain TG1 are deposited respectively under No. I-627 and I-628 at the CNCM.
To reconstitute the complete HIV-2 ROD genome, pROD 35 is linearized with EcoRI and the EcoRI insert of pROD 27-5' is ligated in the correct orientation into this site.
The relationship of HIV-2 to other human and simian retroviruses was surmised from hybridization experiments. The relative homology of the different regions of the HIV-1 and 2 genomes was determined by hybridization of fragments of the cloned HIV-1 genome with the labelled .lambda.ROD 4 expected to contain the complete HIV-2 genome (FIG. 3, B). Even in very low stringency conditions (Tm-42.degree. C.), the hybridization of HIV-1 and 2 was restricted to a fraction of their genomes, principally the qaq gene (dots 1 and 2), the reverse transcriptase domain in pol (dot 3), the end of pol and the Q (or sor) genes (dot 5) and the F gene (or 3' orf) and 3' LTR (dot 11). The HIV-1 fragment used to detect the HIV-2 cDNA clones contained the dot 11 subclone, which hybridized well to HIV-2 under non-stringent conditions. Only the signal from dot 5 persisted after stringent washing. The envelope gene, the region of the tat gene and a part of pol thus seemed very divergent. These data, along with the LTR sequence obtained (FIG. 1, B), indicated that HIV-2 is not an envelope variant of HIV-1, as are African isolates from Zaire described by Alizon et al., Cell 40:63-74 (1986).
It was observed that HIV-2 is related more closely to the Simian Immunodeficiency Virus (SIV) than it is to HIV-1. This correlation has been described by F. Clavel et al. in C. R. Acad. Sci. (Paris) 302: 485-488 (1986) and F. Clavel et al. in Science 233: 343-346 (1986), both of which are specifically incorporated herein by reference. Simian Immunodeficiency Virus (also designated Simian T-cell Lymphotropic Virus Type 3, STLV-3) is a retrovirus first isolated from captive macaques with an AIDS-like disease in the USA. This simian virus has been described by M. D. Daniel et al. in Science 228: 1201-1204 (1985), specifically incorporated herein by reference.
All the SIV proteins, including the envelope, are immune precipitated by sera from HIV-2 infected patients, whereas the serological cross-reactivity of HIV-1 to 2 is restricted to the core proteins. However SIV and HIV-2 can be distinguished by slight differences in the apparent molecular weight of their proteins.
In terms of nucleotide sequence, it also appears that HIV-2 is closely related to SIV. The genomic RNA of SIV can be detected in stringent conditions as shown in FIG. 2, C by HIV-2 probes corresponding to the LTR and 3' end of the genome (E2) or to the gag or pol genes. Under the same conditions, HIV-1 derived probes do not detect the SIV genome as shown in FIG. 2, D.
In Southern blots of DNA from SIV-infected cells, a restriction pattern clearly different from HIV-2.sub.ROD and other isolates is seen. All the bands persist after a stringent washing, even though the signal is considerably weakened, indicating a sequence homology throughout the genomes of HIV-2 and SIV. It has recently been shown that baboons and macaques could be infected experimentally by HIV-2, thereby providing an interesting animal model for the study of the HIV infection and its preventive therapy. Indeed, attempts to infect non-human primates with HIV-1 have been successful only in chimpanzees, which are not a convenient model.
From an initial survey of the restriction maps for certain of the HIV-2 isolates obtained according to the methods described herein, it is already apparent that HIV-2, like HIV-1, undergoes restriction site polymorphism. FIG. 4 A depicts examples of such differences for three isolates, all different one from another and from the cloned HIV-2.sub.ROD. it is very likely that these differences at the nucleotide level are accompanied by variations in the amino-acid sequence of the viral proteins, as evidenced in the case of HIV-1 and described by M. Alizon et al. in Cell 46: 63-74 (1986), specifically incorporated herein by reference. It is also to be expected that the various isolates of HIV-2 will exhibit amino acid heterogeneities. See, for example, Clavel et al., Nature 324 (18):691-695 (1986), specifically incorporated herein by reference.
Further, the characterization of HIV-2 will also delineate the domain of the envelope glycoprotein that is responsible for the binding of the surface of the target cells and the subsequent internalization of the virus. This interaction was shown to be mediated by the CD4 molecule itself in the case of HIV-1 and similar studies tend to indicate that HIV-2 uses the same receptor. Thus, although there is wide divergence between the env genes of HIV-1 and 2, small homologous domains of the envelopes of the two HIV could represent a candidate receptor binding site. This site could be used to raise a protective immune response against this group of retroviruses.
From the data discussed herein, certain nucleotide sequences have been identified which are capable of being used as probes in diagnostic methods to obtain the immunological reagents necessary to diagnose an HIV-2 infection. In particular, these sequences may be used as probes in hybridization reactions with the genetic material of infected patients to indicate whether the RNA of the HIV-2 virus is present in these patient's lymphocytes or whether an analogous DNA is present. In this embodiment, the test methods which may be utilized include Northern blots, Southern blots and dot blots. One particular nucleotide sequence which may be useful as a probe is the combination of the 5 kb. HindIII fragment of ROD 4 and the E2 cONA used in FIG. 4.
In addition, the genetic sequences of the HIV-2 virus may be used to create the polypeptides encoded by these sequences. Specifically, these polypeptides may be created by expression of the cDNA obtained according to the teachings herein in hosts such as bacteria, yeast or animal cells. These polypeptides may be used in diagnostic tests such as immunofluorescence assays (IFA), radioimmunoassays (RIA) and Western Blot tests.
Moreover, it is also contemplated that additional diagnostic tests, including additional immunodiagnostic tests, may be developed in which the DNA probes or the polypeptides of this invention may serve as one of the diagnostic reagents. The invention described herein includes these additional test methods.
In addition, monoclonal antibodies to these polypeptides or fragments thereof may be created. The monoclonal antibodies may be used in immunodiagnostic tests in an analogous manner as the polypeptides described above.
The polypeptides of the present invention may also be used as immunogenic reagents to induce protection against infection by HIV-2 viruses. In this embodiment, the polypeptides produced recombinant-DNA techniques would function as vaccine agents.
Also, the polypeptides of this invention may be used in competitive assays to test the ability of various antiviral agents to determine their ability to prevent the virus from fixing on its target.
Thus, it is to be understood that application of the teachings of the present invention to a specific problem or environment will be within the capabilities of one having ordinary skill in the art in light of the teachings contained herein. Examples of the products of the present invention and representative processes for their isolation and manufacture appear above and in the following examples.
EXAMPLES
Example 1: Cloning of a cDNA Complementary to Genomic RNA From HIV-2 Virions
HIV-2 virions were purified from 5 liters of supernatant from a culture of the CEM cell line infected with the ROD isolate and a cDNA first strand using oligo (dT) primer was synthesized in detergent activated endogenous reaction on pelleted virus, as described by M. Alizon et al. in Nature, 312: 757-760 (1984), specifically incorporated herein by reference. RNA-cDNA hybrids were purified by phenol-chloroform extraction and ethanol precipitation. The second-strand cDNA was created by the DNA polymerase I/RNAase H method of Gubler and Hoffman in Gene, 25: 263-269 (1983), specifically incorporated herein by reference, using a commercial cDNA synthesis kit obtained from Amersham. After attachment of EcoRI linkers (obtained from Pharmacia), EcoRI digestion, and ligation into EcoRI-digested dephosphorylated M13 tg 130 vector (obtained from Amersham), a cDNA library was obtained by transformation of the E. coli TGl strain. Recombinant plaques (10.sup.4) were screened in situ on replica filters with the 1.5 kb. HindIII fragment from clone J19, corresponding to the 3' part of the genome of the LAV.sub.BRU isolate of HIV-1, .sup.32 P labelled to a specific activity of 10.sup.9 cpm .mu.g. The filters were prehybridized in 5.times.SSC, 5.times. Denhardt solution, 25% formamide, and denatured salmon sperm DNA (100 .mu.g/ml.) at 37.degree. C. for 4 hours and hybridized for 16 hours in the same buffer (Tm -42.degree. C.) plus 4.times.10.sup.7 cpm of the labelled probe (10.sup.6 cpm/ml. of hybridization buffer). The washing was done in 5.times.SSC, 0.1% SDS at 25.degree. C. for 2 hours. 20.times.SSC is 3M NaCl, 0.3M Na citrate. Positive plaques were purified and single-stranded M13 DNA prepared and end-sequenced according to the method described in Proc. Nat'l. Acad. Sci. USA, 74: 5463-5467 (1977) of Sanger et al.
Example 2: Hybridization of DNA from HIV-1 and HIV-2 Infected Cells and RNA from HIV-1 and 2 and SIV Virons With a Probe Derived From an HIV-2 Cloned CDNA
DNA was extracted from infected CEM cells continuously producing HIV-1 or 2. The DNA digested with 20 .mu.g of PstI or undigested, was electrophoresed on a 0.8% agarose gel, and Southern-transferred to nylon membrane. Virion dot-blots were prepared in duplicate, as described by F. Clavel et al. in Science 233: 343-346 (1986), specifically incorporated herein by reference, by pelleting volumes of supernatant corresponding to the same amount of reverse transcriptase activity. Prehybridization was done in 50% formamide, 5.times.SSC, 5.times.Denhard solution, and 100 mg./ml. denatured salmon sperm DNA for 4 hours at 42.degree. C. Hybridization was performed in the same buffer plus 10% Dextran sulphate, and 10.sup.6 cpm/ml. of the labelled E2 insert (specific activity 10.sup.9 cpm/.mu.g.) for 16 hours at 42.degree. C. Washing was in 0.1.times.SSC, 0.1% SDS for 2.times.30 mn. After exposition for 16 hours with intensifying screens, the Southern blot was dehybridized in 0.4 N NaOH, neutralized, and rehybridized in the same conditions to the HIV-1 probe labelled to 10.sup.9 cpm/.mu.g.
Example 3: Cloning in Lambda Phage of the Complete Provirus DNA of HIV-2
DNA from the HIV-2.sub.ROD infected CEM (FIG. 2, lanes a and c) was partially digested with Sau3AI. The 9-15 kb. fraction was selected on a 5-40% sucrose gradient and ligated to BamHI arms of the lambda L47.1 vector. Plaques (2.times.10.sup.6) obtained after in vitro packaging and plating on E. coli LA 101 strain were screened in situ with the insert from the E2 cDNA clone. Approximately 10 positive clones were plaque purified and propagated on E. coli C600 recBC. The ROD 4, 27, and 35 clones were amplified and their DNA characterized by restriction mapping and Southern blotting with the HIV-2 cDNA clone under stringent conditions, and gag-pol probes from HIV-1 used under non stringent conditions.
Example 4: Complete Genomic Sequence of the ROD HIV-2 Isolate
Experimental analysis of the HIV-2 ROD isolate yielded the following sequence which represents the complete genome of this HIV-2 isolate. Genes and major expression products identified within the following sequence are indicated by nucleotides numbered below:
1) GAG gene (546-2111) expresses a protein product having a molecular weight of around 55 kd and is cleaved into the following proteins:
a) p 16 (546-950)
b) p 26 (951-1640)
c) p 12 (1701-2111)
2) polymerase (1829-4936)
3) Q protein (4869-5513)
4) R protein (5682-5996)
5) X protein (5344-5679)
6) Y protein (5682-5996)
7) Env protein (6147-8720)
8) F protein (8557-9324)
9) TAT gene (5845-6140 and 8307-8400) is expressed by two exons separated by introns.
10) ART protein (6071-6140 and 8307-8536) is similarly the expression product of two exons.
11) LTR:R (1-173 and 9498-9671)
12) U5 (174-299)
13) U3 (8942-9497)
It will be known to one of skill in the art that the absolute numbering which has been adopted is not essential. For example, the nucleotide within the LTR which is designated as "1" is a somewhat arbitrary choice. What is important is the sequence information provided.
GGTCGCTCTGCGGAGAGGCTGGCAGATTGAGCCCTGGGAGGTTCTCTCCAGCACTAGCAG - . . . . . . GTAGAGCCTGGGTGTTCCCTGCTAGACTCTCACCAGCACTTGGCCGGTGCTGGGCAGACG - . . . 100 . . GCCCCACGCTTGCTTGCTTAAAAACCTCTTAATAAAGCTGCCAGTTAGAAGCAAGTTAAG - . . . . . . TGTGTGCTCCCATCTCTCCTAGTCGCCGCCTGGTCATTCGGTGTTCACCTGAGTAACAAG - . 200 . . . . ACCCTGGTCTGTTAGGACCCTTCTTGCTTTGGGAAACCGAGGCAGGAAAATCCCTAGCAG - . . . . . 300 GTTGGCGCCTGAACAGGGACTTGAAGAAGACTGAGAAGTCTTGGAACACGGCTGACTGAA - . . . . . . GGCAGTAAGGGCGGCAGGAACAAACCACGACGGAGTGCTCCTAGAAAGGCGCGGGCCGAG - . . . 400 . . GTACCAAAGGCAGCGTGTGGAGCGGGAGGAGAAGAGGCCTCCGGGTGAAGGTAAGTACCT - . . . . . . ACACCAAAAACTGTAGCCGAAAGGGCTTGCTATCCTACCTTTAGACAGGTAGAAGATTGT - . 500 . . . . MetGlyAlaArgAsnSerValLeuArgGlyLysLysAlaAspGluLeuGluArgIle GGGAGATGGGCGCGAGAAACTCCGTCTTGAGAGGGAAAAAAGCAGATGAATTAGAAAGAA - . . . . . 600 ArgLeuArgProGlyGlyLysLysLysTyrArgLeuLysHisIleValTrpAlaAlaAsn TCAGGTTACGGCCCGGCGGAAAGAAAAAGTACAGGCTAAAACATATTGTGTGGGCAGCGA - . . . . . . LysLeuAspArgPheGlyLeuAlaGluSerLeuLeuGluSerLysGluGlyCysGlnLys ATAAATTGGACAGATTCGGATTACCAGAGAGCCTGTTGGAGTCAAAAGAGGGTTGTCAAA - . . . 700 . . IleLeuThrValLeuAspProMetValProThrGlySerGluAsnLeuLysSerLeuPhe AAATTCTTACAGTTTTAGATCCAATGGTACCGACAGGTTCAGAAAATTTAAAAAGTCTTT - . . . . . . AsnThrValCysValIleTrpCysIleHisAlaGluGluLysValLysAspThrGluGly TTAATACTGTCTGCGTCATTTGGTGCATACACGCAGAAGAGAAAGTGAAAGATACTGAAG - . 800 . . . . AlaLysGlnIleValArgArgHisLeuValAlaGluThrGlyThrAlaGluLysMetPro GAGCAAAACAAATAGTGCGGAGACATCTAGTGGCAGAAACAGGAACTGCAGAGAAAATGC - . . . . . 900 SerThrSerArgProThrAlaProSerSerGluLysGlyGlyAsnTyrProValGlnHis CAAGCACAAGTAGACCAACAGCACCATCTAGCGAGAAGGGAGGAAATTACCCAGTGCAAC - . . . . . . ValGlyGlyAsnTyrThrHisIleProLeuSerProArgThrLeuAsnAlaTrpValLys ATGTAGGCGGCAACTACACCCATATACCGCTGAGTCCCCGAACCCTAAATGCCTGGGTAA - . . . 1000 . . LeuValGluGluLysLysPheGlyAlaGluValValProGlyPheGlnAlaLeuSerGlu AATTAGTAGAGGAAAAAAAGTTCGGGGCAGAAGTAGTGCCAGGATTTCAGGCACTCTCAG - . . . . . . GlyCysThrProTyrAspIleAsnGlnMetLeuAsnCysValGlyAspHisGlnAlaAla AAGGCTGCACGCCCTATGATATCAACCAAATGCTTAATTGTGTGGGCGACCATCAAGCAG - . 1100 . . . . MetGlnIleIleArgGluIleIleAsnGluGluAlaAlaGluTrpAspValGlnLisPro CCATGCAGATAATCAGGGAGATTATCAATGAGGAAGCAGCAGAATGGGATGTGCAACATC - . . . . . 1200 IleProGlyProLeuProAlaGlyGlnLeuArgGluProArgGlySerAspIleAlaGly CAATACCAGGCCCCTTACCAGCGGGGCAGCTTAGAGAGCCAAGGGGATCTGACATAGCAG - . . . . . . ThrThrSerThrValGluGluGlnIleGlnTrpMetPheArgProGlnAsnProValPro CGACAACAAGCACAGTAGAAGAACAGATCCAGTGGATGTTTAGGCCACAAAATCCTGTAC - . . . 1300 . . ValGlyAsnIleTyrArgArgTrpIleGlnIleGlyLeuGlnLysCysValArgMetTyr CAGTAGGAAACATCTATAGAAGATGGATCCAGATAGGATTGCAGAAGTGTGTCAGGATGT - . . . . . . AsnProThrAsnIleLeuAspIleLysGlnGlyProLysGluProPheGlnSerTyrVal ACAACCCGACCAACATCCTAGACATAAAACAGGGACCAAAGGAGCCGTTCCAAAGCTATG - . 1400 . . . . AspArgPheTyrLysSerLeuArgAlaGluGlnThrAspProAlaValLysAsnTrpMet TAGATAGATTCTACAAAAGCTTGAGGGCAGAACAAACAGATCCAGCAGTGAAGAATTGGA - . . . . . 1500 ThrGlnThrLeuLeuValGlnAsnAlaAsnProAspCysLysLeuValLeuLysGlyLeu TGACCCAAACACTGCTAGTACAAAATGCCAACCCAGACTGTAAATTAGTGCTAAAAGGAC - . . . . . . GlyMetAsnProThrLeuGluGluMetLeuThrAlaCysGlnGlyValGlyGlyProGly TAGGGATGAACCCTACCTTAGAAGAGATGCTGACCGCCTGTCAGGGGGTAGGTGGGCCAG - . . . 1600 . . GlnLysAlaArgLeuMetAlaGluAlaLeuLysGluValIleGlyProAlaProIlePro GCCAGAAAGCTAGATTAATGGCAGAGGCCCTGAAAGAGGTCATAGGACCTGCCCCTATCC - . . . . . . PheAlaAlaAlaGlnGlnArgLysAlaPheLysCysTrpAsnCysGlyLysGluGlyHis CATTCGCAGCAGCCCAGCAGAGAAAGGCATTTAAATGCTGGAACTGTGGAAAGGAAGGGC - . 1700 . . . . SerAlaArgGlnCysArgAlaProArgArgGlnGlyCysTrpLysCysGlyLysProGly ACTCGGCAAGACAATGCCGAGCACCTAGAAGGCAGGGCTGCTGGAAGTGTGGTAAGCCAG - . . . . . 1800 ThrGlyArgPhePheArgThrGlyProLeuGly HisIleMetThrAsnCysProAspArgGlnAlaGlyPheLeuGlyLeuGlyProTrpGly GACACATCATGAGAAACTGCCCAGATAGACAGGCAGGTTTTTTAGGACTGGGCCCTTGGG - . . . . . . LysGluAlaProGlnLeuProArgGlyProSerSerAlaGlyAlaAspThrAsnSerThr LysLysProArgAsnPheProValAlaGlnValProGlnGlyLeuThrProThrAlaPro GAAAGAAGCCGCGCAACTTCCCCGTGGCCCAAGTTCCGCAGGGGCTGACACCAACAGCAC - . . . 1900 . . ProSerGlySerSerSerGlySerThrGlyGluIleTyrAlaAlaArgGluLysThrGlu ProValAspProAlaValAspLeuLeuGluLysTyrMetGlnGlnGlyLysArgGlnArg CCCCAGTGGATCCAGCAGTGGATCTACTGGAGAAATATATGCAGCAAGGGAAAAGACAGA - . . . . . . ArgAlaGluArgGluThrIleGlnGlySerAspArgGlyLeuThrAlaProArgAlaGly GluGlnArgGluArgProTyrLysGluValThrGluAspLeuLeuHisLeuGluGlnGly GAGAGCAGAGAGAGAGACCATACAAGGAAGTGACAGAGGACTTACTGCACCTCGAGCAGG - . 2000 . . . . GlyAspThrIleGlnGlyAlaThrAsnArgGlyLeuAlaAlaProGlnPheSerLeuTrp GluThrProTyrArgGluProProThrGluAspLeuLeuHisLeuAsnSerLeuPheGly GGGAGACACCATACAGGGAGCCACCAACAGAGGACTTGCTGCACCTCAATTCTCTCTTTG - . . . . . 2100 LysArgProValValThrAlaTyrIleGluGlyGlnProValGluValLeuLeuAspThr LysAspGln GAAAAGACCAGTAGTCACAGCATACATTGAGGGTCAGCCAGTAGAAGTCTTGTTAGACAC - . . . . . . GlyAlaAspAspSerIleValAlaGlyIleGluLeuGlyAsnAsnTyrSerProLysIle AGGGGCTGACGACTCAATAGTAGCAGGAATAGAGTTAGGGAACAATTATAGCCCAAAAAT - . . . 2200 . . ValGlyGlyIleGlyGlyPheIleAsnThrLysGluTyrLysAsnValGluIleGluVal AGTAGGGGGAATAGGGGGATTCATAAATACCAAGGAATATAAAAATGTAGAAATAGAAGT - . . . . . . LeuAsnLysLysValArgAlaThrIleMetThrGlyAspThrProIleAsnIlePheGly TCTAAATAAAAAGGTACGGGCCACCATAATGACAGGCGACACCCCAATCAACATTTTTGG - . 2300 . . . . ArgAsnIleLeuThrAlaLeuGlyMetSerLeuAsnLeuProValAlaLysValGluPro CAGAAATATTCTGACAGCCTTAGGCATGTCATTAAATCTACCAGTCGCCAAAGTAGAGCC - . . . . . 2400 IleLysIleMetLeuLysProGlyLysAspGlyProLysLeuArgGlnTrpProLeuThr AATAAAAATAATGCTAAAGCCAGGGAAAGATGGACCAAAACTGAGACAATGGCCCTTAAC - . . . . . . LysGluLysIleGluAlaLeuLysGluIleCysGluLysMetGluLysGluGlyGlnLeu AAAAGAAAAAATAGAAGCACTAAAAGAAATCTGTGAAAAAATGGAAAAAGAAGGCCAGCT - . . . 2500 . . GluGluAlaProProThrAsnProTyrAsnThrProThrPheAlaIleLysLysLysAsp AGAGGAAGCACCTCCAACTAATCCTTATAATACCCCCACATTTGCAATCAAGAAAAAGGA - . . . . . . LysAsnLysTrpArgMetLeuIleAspPheArgGluLeuAsnLysValThrGlnAspPhe CAAAAACAAATGGAGGATGCTAATAGATTTCAGAGAACTAAACAAGGTAACTCAAGATTT - . 2600 . . . . ThrGluIleGlnLeuGlyIleProHisProAlaGlyLeuAlaLysLysArgArgIleThr CACAGAAATTCAGTTAGGAATTCCACACCCAGCAGGGTTGGCCAAGAAGAGAAGAATTAC - . . . . . 2700 ValLeuAspValGlyAspAlaTyrPheSerIleProLeuHisGluAspPheArgProTyr TGTACTAGATGTAGGGGATGCTTACTTTTCCATACCACTACATGAGGACTTTAGACCATA - . . . . . . ThrAlaPheThrLeuProSerValAsnAsnAlaGluProGlyLysArgTyrIleTyrLys TACTGCATTTACTCTACCATCAGTGAACAATGCAGAACCAGGAAAAAGATACATATATAA - . . . 2800 . . ValLeuProGlnGlyTrpLysGlySerProAlaIlePheGlnHisThrMetArgGlnVal AGTCTTGCCACAGGGATGGAAGGGATCACCAGCAATTTTTCAACACACAATGAGACAGGT - . . . . . . LeuGluProPheArgLysAlaAsnLysAspValIleIleIleGlnTyrMetAspAspIle ATTAGAACCATTCAGAAAAGCAAACAAGGATGTCATTATCATTCAGTACATGGATGATAT - . 2900 . . . . LeuIleAlaSerAspArgThrAspLeuGluHisAspArgValValLeuGlnLeuLysGlu CTTAATAGCTAGTGACAGGACAGATTTAGAACATGATAGGGTAGTCCTGCAGCTCAAGGA - . . . . . 3000 LeuLeuAsnGlyLeuGlyPheSerThrProAspGluLysPheGlnLysAspProProTyr ACTTCTAAATGGCCTAGGATTTTCTACCCCAGATGAGAAGTTCCAAAAAGACCCTCCATA - . . . . . . HisTrpMetGlyTyrGluLeuTrpProThrLysTrpLysLeuGlnLysIleGlnLeuPro CCACTGGATGGGCTATGAACTATGGCCAACTAAATGGAAGTTGCAGAAAATACAGTTGCC - . . . 3100 . . GlnLysGluIleTrpThrValAsnAspIleGlnLysLeuValGlyValLeuAsnTrpAla CCAAAAAGAAATATGGACAGTCAATGACATCCAGAAGCTAGTGGGTGTCCTAAATTGGGC - . . . . . . AlaGlnLeuTyrProGlyIleLysThrLysHisLeuCysArgLeuIleArgGlyLysMet AGCACAACTCTACCCAGGGATAAAGACCAAACACTTATGTAGGTTAATCAGAGGAAAAAT - . 3200 . . . . ThrLeuThrGluGluValGlnTrpThrGluLeuAlaGluAlaGluLeuGluGluAsnArg GACACTCACAGAAGAAGTACAGTGGACAGAATTACCAGAAGCAGAGCTAGAAGAAAACAG - . . . . . 3300 IleIleLeuSerGlnGluGlnGluGlyHisTyrTyrGlnGluGluLysGluLeuGluAla AATTATCCTAAGCCAGGAACAAGAGGGACACTATTACCAAGAAGAAAAAGAGCTAGAAGC - . . . . . . ThrValGlnLysAspGlnGluAsnGlnTrpThrTyrLysIleHisGlnGluGluLysIle AACAGTCCAAAAGGATCAAGAGAATCAGTGGACATATAAAATACACCAGGAAGAAAAAAT - . . . 3400 . . LeuLysValGlyLysTyrAlaLysValLysAsnThrHisThrAsnGlyIleArgLeuLeu TCTAAAAGTAGGAAAATATGCAAAGGTGAAAAACACCCATACCAATGGAATCAGATTGTT - . . . . . . AlaGlnValValGlnLysIleGlyLysGluAlaLeuValIleTrpGlyArgIleProLys AGCACAGGTAGTTCAGAAAATAGGAAAAGAAGCACTAGTCATTTGGGGACCAATACCAAA - . 3500 . . . . PheHisLeuProValGluArgGluIleTrpGluGlnTrpTrpAspAsnTyrTrpGlnVal ATTTCACCTACCAGTAGAGAGAGAAATCTGGGAGCAGTGGTGGGATAACTACTGGCAAGT - . . . . . 3600 ThrTrpIleProAspTrpAspPheValSerThrProProLeuValArgLeuAlaPheAsn GACATGGATCCCAGACTGGGACTTCGTGTCTACCCCACCACTGGTCAGGTTAGCGTTTAA - . . . . . . LeuValGlyAspProIleProGlyAlaGluThrPheTyrThrAspGlySerCysAsnArg CCTGGTAGGGGATCCTATACCAGGTGCAGAGACCTTCTACACAGATCGATCCTGCAATAG - . . . 3700 . . GlnSerLysGluGlyLysAlaGlyTyrValThrAspArgGlyLysAspLysValLysLys GCAATCAAAAGAAGGAAAAGCAGGATATGTAACAGATAGAGGGAAAGACAAGGTAAAGAA - . . . . . . LeuGluGlnThrThrAsnGlnGlnAlaGluLeuGluAlaPheAlaMetAlaLeuThrAsp ACTAGAGCAAACTACCAATCAGCAAGCAGAACTAGAAGCCTTTGCGATGGCACTAACAGA - . 3800 . . . . SerGlyProLysValAsnIleIleValAspSerGlnTyrValMetGlyIleSerAlaSer CTCGGGTCCAAAAGTTAATATTATAGTAGACTCACAGTATGTAATGGGGATCAGTGCAAG - . . . . . 3900 GlnProThrGluSerGluSerLysIleValAsnGlnIleIleGluGluMetIleLysLys CCAACCAACAGAGTCAGAAAGTAAAATAGTGAACCAGATCATAGAAGAAATGATAAAAAA - . . . . . . GluAlaIleTyrValAlaTrpValProAlaHisLysGlyIleGlyGlyAsnGlnGluVal GGAAGCAATCTATGTTGCATGGGTCCCAGCCCACAAAGGCATAGGGGGAAACCAGGAAGT - . . . 4000 . . AspHisLeuValSerGlnGlyIleArgGlnValLeuPheLeuGluLysIleGluProAla AGATCATTTAGTGAGTCAGGGTATCAGACAAGTGTTGTTCCTGGAAAAAATAGAGCCCGC - . . . . . . GlnGluGluHisGluLysTyrHisSerAsnValLysGluLeuSerHisLysPheGlyIle TCAGGAAGAACATGAAAAATATCATAGCAATGTAAAAGAACTGTCTCATAAATTTGGAAT - . 4100 . . . . ProAsnLeuValAlaArgGlnIleValAsnSerCysAlaGlnCysGlnGlnLysGlyGlu ACCCAATTTAGTGGCAAGGCAAATAGTAAACTCATGTGCCCAATGTCAACAGAAAGGGGA - . . . . . 4200 AlaIleHisGlyGlnValAsnAlaGluLeuGlyThrTrpGlnMetAspCysThrHisLeu AGCTATACATGGGCAAGTAAATGCAGAACTAGCCACTTGGCAAATGGACTGCACACATTT - . . . . . . GluGlyLysIleIleIleValAlaValHisValAlaSerGlyPheIleGluAlaGluVal AGAAGGAAAGATCATTATAGTAGCAGTACATGTTGCAAGTGGATTTATAGAAGCAGAAGT - . . . 4300 . . IleProGlnGluSerGlyArgGlnThrAlaLeuPheLeuLeuLysLeuAlaSerArgTrp CATCGCACAGGAATCAGGAAGACAAACAGCACTCTTCCTATTGAAACTGGCAAGTAGGTG - . . . . . . ProIleThrHisLeuHisThrAspAsnGlyAlaAsnPheThrSerGlnGluValLysMet GCCAATAACACACTTGCATACAGATAATGGTGCCAACTTCACTTCACAGGAGGTGAAGAT - . 4400 . . . . ValAlaTrpTrpIleGlyIleGluGlnSerPheGlyValProTyrAsnProGlnSerGln GGTAGCATGGTGGATAGGTATAGAACAATCCTTTGGAGTACCTTACAATCCACAGAGCCA - . . . . . 4500 GlyValValGluAlaMetAsnHisHisLeuLysAsnGluIleSerArgIleArgGluGln AGGAGTAGTAGAAGCAATGAATCACCATCTAAAAAACCAAATAAGTAGAATCAGAGAACA - . . . . . . AlaAsnThrIleGluThrIleValLeuMetAlaIleHisCysMetAsnPheLysArgArg GGCAAATACAATAGAAACAATAGTACTAATGGCAATTCATTGCATGAATTTTAAAAGAAG - . . . 4600 . . GlyGlyIleGlyAspMetThrProSerGluArgLeuIleAsnMetIleThrThrGluGln GGGGGGAATAGGGGATATGACTCCATCAGAAAGATTAATCAATATGATCACCACAGAACA - . . . . . . GluIleGlnPheLeuGlnAlaLysAsnSerLysLeuLysAspPheArgValTyrPheArg AGAGATACAATTCCTCCAAGCCAAAAATTCAAAATTAAAAGATTTTCGGGTCTATTTCAG - . 4700 . . . . GluGlyArgAspGlnLeuTrpLysGlyProGlyGluLeuLeuTrpLysGlyGluGlyAla AGAAGGCAGACATCAGTTGTGGAAAGGACCTGGGGAACTACTGTGGAAAGGAGAAGGAGC - . . . . . 4800 ValLeuValLysValGlyThrAspIleLysIleIleProArgArgLysAlaLysIleIle AGTCCTAGTCAAGGTAGGAACAGACATAAAAATAATACCAAGAAGGAAAGCCAAGATCAT - . . . . . . ArgAspTyrGlyGlyArgGlnGluMetAspSerGlySerHisLeuGluGlyAlaArgGlu MetGluGluAspLysArgTrpIleValValProThrTrpArgValProGlyArg CAGACACTATGGAGGAAGACAAGAGATGGATAGTGGTTCCCACCTGGAGGGTGCCAGGGA - . . . 4900 . . AspGlyGluMetAla MetGluLysTrpHisSerLeuValLysTyrLeuLysTyrLysThrLysAspLeuGluLys GGATGGAGAAATGGCATAGCCTTGTCAAGTATCTAAAATACAAAACAAAGGATCTAGAAA - . . . . . . ValCysTyrValProHisHisLysValGlyTrpAlaTrpTrpThrCysSerArgValIle AGGTGTGCTATGTTCCCCACCATAAGGTGGGATGGGCATGGTGGACTTGCAGCAGGGTAA - . 5000 . . . . PheProLeuLysGlyAsnSerHisLeuGluIleGlnAlaTyrTrpAsnLeuThrProGlu TATTCCCATTAAAAGGAAACAGTCATCTAGAGATACAGGCATATTGGAACTTAACACCAG - . . . . . 5100 LysGlyTrpLeuSerSerTyrSerValArgIleThrTrpTyrThrGluLysPheTrpThr AAAAAGGATGGCTCTCCTCTTATTCAGTAAGAATAACTTGGTACACAGAAAAGTTCTGGA - . . . . . . AspValThrProAspCysAlaAspValLeuIleHisSerThrTyrPheProCysPheThr CAGATGTTACCCCAGACTGTGCAGATGTCCTAATACATAGCACTTATTTCCCTTGCTTTA - . . . 5200 . . AlaGlyGluValArgArgAlaIleArgGlyGluLysLeuLeuSerCysCysAsnTyrPro CAGCAGGTGAAGTAAGAAGAGCCATCAGAGGGGAAAAGTTATTGTCCTGCTGCAATTATC - . . . . . . ArgAlaHisArgAlaGlnValProSerLeuGlnPheLeuAlaLeuValValValGlnGln CCCGAGCTCATAGACCCCAGGTACCGTCACTTCAATTTCTGGCCTTAGTGGTAGTCCAAC - . 5300 . . . . MetThrAspProArgGluThrValProProGlyAsnSerGlyGluGluThrIleGly AsnAspArgProGlnArgAspSerThrThrArgLysGlnArgArgArgAspTyrArgArg AAAATGACAGACCCCAGAGAGACAGTACCACCAGGAAACAGCGGCGAAGAGACTATCGGA - . . . . . 5400 GluAlaPheAlaTrpLeuAsnArgThrValGluAlaIleAsnArgGluAlaValAsnHis GlyLeuArgLeuAlaLysGlnAspSerArgSerHisLysGlnArgSerSerGluSerPro GAGGCCTTCGCCTGGCTAAACAGGACAGTAGAAGCCATAAACAGAGAAGCAGTGAATCAC - . . . . . . LeuProArgGluLeuIlePheGlnValTrpGlnArgSerTrpArgTyrTrpHisAspGlu ThrProArgThrTyrPheProGlyValAlaGluValLeuGluIleLeuAla CTACCCCGAGAACTTATTTTCCAGGTGTGGCAGAGGTCCTGGAGATACTGGCATGATGAA - . . . 5500 . . GlnGlyMetSerGluSerTyrThrLysTyrArgTyrLeuCysIleIleGlnLysAlaVal CAAGGGATGTCAGAAAGTTACACAAAGTATAGATATTTGTGCATAATACAGAAAGCAGTG - . . . . . . TyrMetHisValArgLysGlyCysThrCysLeuGlyArgGlyHisGlyProGlyGlyTrp TACATGCATGTTAGGAAAGGGTGTACTTGCCTGGGGAGGGGACATGGGCCAGGAGGGTGG - . 5600 . . . . ArgProGlyProProProProProProProGlyLeuVal MetAlaGluAlaProThrGlu AGACCAGGGCCTCCTCCTCCTCCCCCTCCAGGTCTGGTCTAATGGCTGAAGCACCAACAG - . . . . . 5700 LeuProProValAspGlyThrProLeuArgGluProGlyAspGluTrpIleIleGluIle AGCTCCCCCCGGTGGATGGGACCCCACTGAGGGAGCCAGGGGATGAGTGGATAATAGAAA - . . . . . . LeuArgGluIleLysGluGluAlaLeuLysHisPheAspProArgLeuLeuIleAlaLeu TCTTGAGAGAAATAAAAGAAGAAGCTTTAAAGCATTTTGACCCTCGCTTGCTAATTGCGC - . . . 5800 . . MetGluThrProLeuLysAlaProGluSerSerLeu GlyLysTyrIleTyrThrArgHisGlyAspThrLeuGluGlyAlaArgGluLeuIleLys TTGGCAAATATATCTATACTAGACATGGAGACACCCTTGAAGGCGCCAGAGAGCTCATTA - . . . . . . LysSerCysAsnGluProPheSerArgThrSerGluGlnAspValAlaThrGlnGluLeu ValLeuGlnArgAlaLeuPheThrHisPheArgAlaGlyCysGlyHisSerArgIleGly AAGTCCTGCAACGAGCCCTTTTCACGCACTTCAGAGCAGGATGTGGCCACTCAAGAATTG - . 5900 . . . . AlaArgGlnGlyGluGluIleLeuSerGlnLeuTyrArgProLeuGluThrCysAsnAsn GlnThrArgGlyGlyAsnProLeuSerAlaIleProThrProArgAsnMetGln GCCAGACAAGGGGAGGAAATCCTCTCTCAGCTATACCGACCCCTAGAAACATGCAATAAC - . . . . . 6000 SerCysTyrCysLysArgCycCysTyrHisCysGlnMetCysPheLeuAsnLysGlyLeu TCATGCTATTGTAAGCGATGCTGCTACCATTGTCAGATGTGTTTTCTAAACAAGGGGCTC - . . . . . . GlyIleCysTyrGluArgLysGlyArgArgArgArgThrProLysLysThrLysThrHis MetAsnGluArgAlaAspGluGluGlyLeuGlnArgLysLeuArgLeuIle GGGATATGTTATGAACGAAAGGGCAGACGAAGAAGGACTCCAAAGAAAACTAAGACTCAT - . . . 6100 . . ProSerProThrProAspLys ArgLeuLeuHisGlnThr MetMetAsnGlnLeuLeuIleAlaIleLeuLeuAla CCGTCTCCTACACCAGACAAGTGAGTATGATGAATCAGCTGCTTATTGCCATTTTATTAG - . . . . . . SerAlaCysLeuValTyrCysThrGlnTyrValThrValPheTyrGlyValProThrTrp CTAGTGCTTGCTTAGTATATTGCACCCAATATGTAACTGTTTTCTATGGCGTACCCACGT - . 6200 . . . . LysAsnAlaThrIleProLeuPheCysAlaThrArgAsnArgAspThrTrpGlyThrIle GGAAAAATGCAACCATTCCCCTCTTTTGTGCAACCAGAAATAGGGATACTTGGGGAACCA - . . . . . 6300 GlnCysLeuProAspAsnAspAspTyrGlnGluIleThrLeuAsnValThrGluAlaPhe TACAGTGCTTGCCTGACAATGATGATTATCAGGAAATAACTTTGAATGTAACAGAGCCTT - . . . . . . AspAlaTrpAsnAsnThrValThrGluGlnAlaIleGluAspValTrpHisLeuPheGlu TTGATGCATGGAATAATACAGTAACAGAACAAGCAATAGAAGATGTCTGGCATCTATTCG - . . . 6400 . . ThrSerIleLysProCysValLysLeuThrProLeuCysValAlaMetLysCysSerSer AGACATCAATAAAACCATGTGTCAAACTAACACCTTTATGTGTAGCAATGAAATGCAGCA - . . . . . . ThrGluSerSerThrGlyAsnAsnThrThrSerLysSerThrSerThrThrThrThrThr GCACAGAGAGCAGCACAGGGAACAACACAACCTCAAAGAGCACAAGCACAACCACAACCA - . 6500 . . . . ProThrAspGlnGluGlnGluIleSerGluAspThrProCysAlaArgAlaAspAsnCys CACCCACAGACCAGGAGCAAGAGATAAGTGAGGATACTCCATGCGCACGCGCAGACAACT - . . . . . 6600 SerGlyLeuGlyGluGluGluThrIleAsnCysGlnPheAsnMetThrGlyLeuGluArg GCTCAGGATTGGGAGAGGAAGAAACGATCAATTGCCAGTTCAATATGACAGGATTAGAAA - . . . . . . AspLysLysLysGlnTyrAsnGluThrTrpTyrSerLysAspValValCysGluThrAsn GAGATAAGAAAAAACAGTATAATGAAACATGGTACTCAAAAGATGTGGTTTGTGAGACAA - . . . 6700 . . AsnSerThrAsnGlnThrGlnCysTyrMetAsnHisCysAsnThrSerValIleThrGlu ATAATAGCACAAATCAGACCCAGTGTTACATGAACCATTGCAACACATCAGTCATCACAG - . . . . . . SerCysAspLysHisTyrTrpAspAlaIleArgPheArgTyrCysAlaProProGlyTyr AATCATGTGACAAGCACTATTGGGATGCTATAAGGTTTAGATACTGTGCACCACCGGGTT - . 6800 . . . . AlaLeuLeuArgCysAsnAspThrAsnTyrSerGlyPheAlaProAsnCysSerLysVal ATGCCCTATTAAGATGTAATGATACCAATTATTCAGGCTTTGCACCCAACTGTTCTAAAG - . . . . . 6900 ValAlaSerThrCysThrArgMetMetGluThrGlnThrSerThrTrpPheGlyPheAsn TAGTAGCTTCTACATGCACCAGGATGATGGAAACGCAAACTTCCACATGGTTTGGCTTTA - . . . . . . GlyThrArgAlaGluAsnArgThrTyrIleTyrTrpHisGlyArgAspAsnArgThrIle ATGGCACTAGAGCAGAGAATAGAACATATATCTATTGGCATGGCAGAGATAATAGAACTA - . . . 7000 . . IleSerLeuAsnLysTyrTyrAsnLeuSerLeuHisCysLysArgProGlyAsnLysThr TCATCAGCTTAAACAAATATTATAATCTCAGTTTGCATTGTAAGAGGCCAGGGAATAAGA - . . . . . . ValLysGlnIleMetLeuMetSerGlyHisValPheHisSerHisTyrGlnProIleAsn CAGTGAAACAAATAATGCTTATGTCAGGACATGTGTTTCACTCCCACTACCAGCCGATCA - . 7100 . . . . LysArgProArgGlnAlaTrpCysTrpPheLysGlyLysTrpLysAspAlaMetGlnGlu ATAAAAGACCCAGACAAGCATGGTGCTGGTTCAAAGGCAAATGGAAAGACGCCATGCAGG - . . . . . 7200 ValLysGluThrLeuAlaLysHisProArgTyrArgGlyThrAsnAspThrArgAsnIle AGGTGAAGGAAACCCTTGCAAAACATCCCAGGTATAGAGGAACCAATGACACAAGGAATA - . . . . . . SerPheAlaAlaProGlyLysGlySerAspProGluValAlaTyrMetTrpThrAsnCys TTAGCTTTGCAGCGCCAGGAAAAGGCTCAGACCCAGAAGTAGCATACATGTGGACTAACT - . . . 7300 . . ArgGlyGluPheLeuTyrCysAsnMetThrTrpPheLeuAsnTrpIleGluAsnLysThr GCAGAGGAGAGTTTCTCTACTGCAACATGACTTGGTTCCTCAATTGGATAGAGAATAAGA - . . . . . . HisArgAsnTyrAlaProCysHisIleLysGlnIleIleAsnThrTrpHisLysValGly CACACCGCAATTATGCACCGTGCCATATAAAGCAAATAATTAACACATGGCATAAGGTAG - . 7400 . . . . ArgAsnValTyrLeuProProArgGluGlyGluLeuSerCysAsnSerThrValThrSer GGAGAAATGTATATTTGCCTCCCAGGGAAGGGGAGCTGTCCTGCAACTCAACAGTAACCA - . . . . . 7500 IleIleAlaAsnIleAspTrpGlnAsnAsnAsnGlnThrAsnIleThrPheSerAlaGlu GCATAATTGCTAACATTGACTGGCAAAACAATAATCAGACAAACATTACCTTTAGTGCAG - . . . . . . ValAlaGluLeuTyrArgLeuGluLeuGlyAspTyrLysLeuValGluIleThrProIle AGGTGGCAGAACTATACAGATTGGAGTTGGGAGATTATAAATTGGTAGAAATAACACCAA - . . . 7600 . . GlyPheAlaProThrLysGluLysArgTyrSerSerAlaHisGlyArgHisThrArgGly TTGGCTTCGCACCTACAAAAGAAAAAAGATACTCCTCTGCTCACGGGAGACATACAAGAG - . . . . . . ValPheValLeuGlyPheLeuGlyPheLeuAlaThrAlaGlySerAlaMetGlyAlaAla GTGTGTTCGTGCTAGGGTTCTTGGGTTTTCTCGCAACAGCAGGTTCTGCAATGGGCGCGG - . 7700 . . . . SerLeuThrValSerAlaGlnSerArgThrLeuLeuAlaGlyIleValGlnGlnGlnGln CGTCCCTGACCGTGTCGGCTCAGTCCCGGACTTTACTGGCCGGGATAGTGCAGCAACAGC - . . . . . 7800 GlnLeuLeuAspValValLysArgGlnGlnGluLeuLeuArgLeuThrValTrpGlyThr AACAGCTGTTGGACGTGGTCAAGAGACAACAAGAACTGTTGCGACTGACCGTCTGGGGAA - . . . . . . LysAsnLeuGlnAlaArgValThrAlaIleGluLysTyrLeuGlnAspGlnAlaArgLeu CGAAAAACCTCCAGGCAACAGTCACTGCTATAGAGAAGTACCTACAGGACCAGGCGCGGC - . . . 7900 . . AsnSerTrpGlyCysAlaPheArgGlnValCysHisThrThrValProTrpValAsnAsp TAAATTCATGGGGATGTGCGTTTAGACAAGTCTGCCACACTACTGTACCATGGGTTAATG - . . . . . . SerLeuAlaProAspTrpAspAsnMetThrTrpGlnGluTrpGluLysGlnValArgTyr ATTCCTTAGCACCTGACTGGGACAATATGACGTGGCAGGAATGGGAAAAACAAGTCCGCT - . 8000 . . . . LeuGluAlaAsnIleSerLysSerLeuGluGlnAlaGlnIleGlnGlnGluLysAsnMet ACCTGGAGGCAAATATCAGTAAAAGTTTAGAACAGGCACAAATTCAGCAAGAGAAAAATA - . . . . . 8100 TyrGluLeuGlnLysLeuAsnSerTrpAspIlePheGlyAsnTrpPheAspLeuThrSer TGTATGAACTACAAAAATTAAATAGCTGGGATATTTTTGGCAATTGGTTTGACTTAACCT - . . . . . . TrpValLysTyrIleGlnTyrGlyValLeuIleIleValAlaValIleAlaLeuArgIle CCTGGGTCAAGTATATTCAATATGGAGTCCTTATAATAGTAGCAGTAATAGCTTTAACAA - . . . 8200 . . ValIleTyrValValGlnMetLeuSerArgLeuArgLysGlyTyrArgProValPheSer TAGTGATATATGTAGTACAAATGTTAAGTAGGCTTAGAAAGGGCTATAGGCCTGTTTTCT - . . . . . . SerIleSerThrArgThrGlyAspSerGlnPro AsnProTyrProGlnGlyProGlyThrAlaSerGln SerProProGlyTyrIleGlnGlnIleHisIleHisLysAspArgGlyGlnProAlaAsn CTTCCCCCCCCGGTTATATCCAACAGATCCATATCCACAAGGACCGGGGACAGCCAGCCA - . 8300 . . . . ThrLysLysGlnLysLysThrValGluAlaThrValGluThrAspThrGlyProGlyArg ArgArgAsnArgArgArgArgTrpLysGlnArgTrpArgGlnIleLeuAlaLeuAlaAsp GluGluThrGluGluAspGlyGlySerAsnGlyGlyAspArgTyrTrpProTrpProIle ACGAAGAAACAGAAGAAGACGGTGGAAGCAACGGTGGAGACAGATACTGGCCCTGGCCGA - . . . . . 8400 SerIleTyrThrPheProAspProProAlaAspSerProLeuAspGlnThrIleGlnHis AlaTyrIleHisPheLeuIleArgGlnLeuIleArgLeuLeuThrArgLeuTyrSerIle TAGCATATATACATTTCCTGATCCGCCAGCTGATTCGCCTCTTGACCAGACTATACAGCA - . . . . . . LeuGlnGlyLeuThrIleGlnGluLeuProAspProProThrHisLeuProGluSerGln CysArgAspLeuLeuSerArgSerPheLeuThrLeuGlnLeuIleTyrGlnAsnLeuArg TCTGCAGGGACTTACTATCCAGGAGCTTCCTGACCCTCCAACTCATCTACCAGAATCTCA - . . . 8500 . . ArgLeuAlaGluThr MetGlyAlaSerGlySerLysLys AspTrpLeuArgLeuArgThrAlaPheLeuGlnTyrGlyCysGluTrpIleGlnGluAla GAGACTGGCTGAGACTTAGAACAGCCTTCTTGCAATATGGGTGCGAGTGGATCCAAGAAG - . . . . . . HisSerArgProProArgGlyLeuGlnGluArgLeuLeuArgAlaArgAlaGlyAlaCys PheGlnAlaAlaAlaArgAlaThrArgGluThrLeuAlaGlyAlaCysArgGlyLeuTrp CATTCCAGGCCGCCGCGAGGGCTACAAGAGAGACTCTTGAGGGCGCGTGCAGGGGCTTCT - . 8600 . . . . GlyGlyTyrTrpAsnGluSerGlyGluGluTyrSerArgPheGlnGluGlySerAspArg ArgValLeuGluArgIleGlyArgGlyIleLeuAlaValProArgArgIleArgGlnGly GCAGCGTATTGGAACGAATCGGGAGGGGAATACTCGCGGTTCCAAGAAGGATCAGACAGC - . . . . . 8700 GluGlnLysSerProSerCysGluGlyArgGlnTyrGlnGlnGlyAspPheMetAsnThr AlaGluIleAlaLeuLeu GAGCAGAAATCGCCCTCCTGTGAGGGACGGCAGTATCAGCAGGGAGACTTTATGAATACT - . . . . . . ProTrpLysAspProAlaAlaGluArgGluLysAsnLeuTyrArgGlnGlnAsnMetAsp CCATGGAAGGACCCAGCAGCAGAAAGGGAGAAAAATTTGTACAGGCAACAAAATATGGAT - . . . 8800 . . AspValAspSerAspAspAspAspGlnValArgValSerValThrProLysValProLeu GATGTAGATTCAGATGATGATGACCAAGTAAGAGTTTCTGTCACACCAAAAGTACCACTA - . . . . . . ArgProMetThrHisArgLeuAlaIleAspMetSerHisLeuIleLysThrArgGlyGly AGACCAATGACACATAGATTGGCAATAGATATGTCACATTTAATAAAAACAAGGGGGGGA - . 8900 . . . . LeuGluGlyMetPheTyrSerGluArgArgHisLysIleLeuAsnIleTyrLeuGluLys CTGGAAGGGATGTTTTACAGTGAAAGAAGACATAAAATCTTAAATATATACTTAGAAAAG - . . . . . 9000 GluGluGlyIleIleAlaAspTrpGlnAsnTyrThrHisGlyProGlyValArgTyrPro CAAGAAGGGATAATTGCAGATTGGCAGAACTACACTCATGGGCCAGGAGTAAGATACCCA - . . . . . . MetPhePheGlyTrpLeuTrpLysLeuValProValAspValProGlnGluGlyGluAsp ATGTTCTTTGGGTGGCTATGGAAGCTAGTACCAGTAGATGTCCCACAAGAAGGGGAGGAC - . . . 9100 . . ThrGluThrHisCysLeuValHisProAlaGlnThrSerLysPheAspAspProHisGly ACTGAGACTCACTGCTTAGTACATCCAGCACAAACAAGCAAGTTTGATGACCCGCATGGG - . . . . . . GluThrLeuValTrpGluPheAspProLeuLeuAlaTyrSerTyrGluAlaPheIleArg GAGACACTAGTCTGGGAGTTTGATCCCTTGCTGGCTTATAGTTACGAGGCTTTTATTCGG - . 9200 . . . . TyrProGluGluPheGlyHisLysSerGlyLeuProGluGluGluTrpLysAlaArgLeu TACCCAGAGGAATTTGGGCACAAGTCAGGCCTGCCAGAGGAAGAGTGGAAGGCGAGACTG - . . . . . 9300 LysAlaArgGlyIleProPheSer AAAGCAAGAGCAATACCATTTAGTTAAAGACAGGAACAGCTATACTTGGTCAGGGCAGGA - . . . . . . AGTAACTAACAGAAACAGCTGAGACTGCAGGGACTTTCCAGAAGGGGCTGTAACCAAGGG - . . . 9400 . . AGGGACATGGGAGGAGCTGGTGGGGAACGCCCTCATATTCTCTGTATAAATATACCCGCT - . . . . . . AGCTTGCATTGTACTTCGGTCGCTCTGCGGAGAGGCTGGCAGATTGAGCCCTGGGAGGTT - . 9500 . . . . CTCTCCAGCAGTAGCAGGTAGAGCCTGGGTGTTCCCTGCTAGACTCTCACCAGCACTTGG - . . . . . 9600 CCGGTGCTGGGCAGACGGCCCCACGCTTGCTTGCTTAAAAACCTCCTTAATAAAGCTGCC - . . . . . . AGTTAGAAGCA - .
Example 5: Sequences of the coding Regions for the Envelope Protein and GAG Product of the ROD HIV-2 Isolate
Through experimental analysis of the HIV-2 ROD isolate, the following sequences were identified for the regions encoding the env and gag gene products. One of ordinary skill in the art will recognize that the numbering for both gene regions which follow begins for convenience with "1" rather than the corresponding number for its initial nucleotide as given in Example 4, above, in the context of the complete genomic sequence.
Envelope sequence
Envelope sequence - - MetMetAsnGlnLeuLeuIleAlaIleLeuLeuAlaSerAlaCys ATGATGATCCAGCTGCTTATTGCCATTTTATTAGCTAGTGCTTGC . . . . LeuValTyrCysThrGlnTyrValThrValPheTyrGlyValPro TTAGTATATTGCACCCAATATGTAACTGTTTTCTATGGCGTACCC . . . . . ThrTrpLysAsnAlaThrIleProLeuPheCysAlaThrArgAsn ACGTGGAAAAATGCAACCATTCCCCTGTTTTGTGCAACCAGAAAT 100 . . . ArgAspThrTrpGlyThrIleGlnCysLeuProAspAsnAspAsp AGGGATACTTGGGGAACCATACAGTGCTTGCCTGACAATGATGAT . . . . . TyrGlnGluIleThrLeuAsnValThrGluAlaPheAspAlaTrp TATCAGGAAATAACTTTGAATGTAACAGAGGCTTTTGATGCATGG . 200 . . AsnAsnThrValThrGluGlnAlaIleGluAspValTrpHisLeu AATAATACAGTAACAGAACAAGCAATAGAAGATGTCTGGCATCTA . . . . . PheGluThrSerIleLysProCysValLysLeuThrProLeuCys TTCGAGACATCAATAAAACCATGTGTGAAACTAACACCTTTATGT . . 300 . ValAlaMetLysCysSerSerThrGluSerSerThrGlyAsnAsn GTAGCAATGAAATGCAGCAGCACAGAGAGCAGCACAGGGAACAAC . . . . . ThrThrSerLysSerThrSerThrThrThrThrThrProThrAsp ACAACCTCAAAGAGCACAAGCACAACCACAACCACACCCAGAGAG . . . 400 GlnGluGlnGluIleSerGluAspThrProCysAlaArgAlaAsp CAGGAGCAAGAGATAAGTGAGGATACTCCATGCGCACGCGCAGAC . . . . . AsnCysSerGlyLeuGlyGluGluGLuThrIleAsnCysGlnPhe AACTGCTCAGGATTGGGAGAGGAAGAAACGATCAATTGCCAGTTC . . . . AsnMetThrGlyLeuGluArgAspLysLysLysGlnTyrAsnGlu AATATGACAGGATTAGAAAGAGATAAGAAAAAACAGTATAATGAA 500 . . . . ThrTrpTyrSerLysAspValValCysGluThrAsnAsnSerThr ACATGGTACTCAAAAGATGTGGTTTGTGAGACAAATAATAGCACA . . . . AsnGlnThrGlnCysTyrMetAsnHisCysAsnThrSerValIle AATCAGACCCAGTGTTACATGAACCATTGCAACACATCAGTCATC . 600 . . . ThrGluSerCysAspLysHisTyrTrpAspAlaIleArgPheArg ACAGAATCATGTGACAAGCACTATTGGGATGCTATAAGGTTTAGA . . . . TyrCysAlaProProGlyTyrAlaLeuLeuArgCysAsnAspThr TACTGTGCACCACCGGGTTATGCCCTATTAAGATGTAATGATACC . . 700 . . AsnTrySerGlyPheAlaProAsnCysSerLysValValAlaSer AATTATTCAGGCTTTGCACCCAACTGTTCTAAAGTAGTAGCTTCT . . . . ThrCysThrArgMetMetGluThrGlnThrSerThrTrpPheGly ACATGCACCAGGATGATGGAAACGCAAACTTCCACATGGTTTGGC . . . 800 . PheAsnGlyThrArgAlaGluAsnArgThrTyrIleTyrTrpHis TTTAATGGCACTAGAGCAGAGAATAGAACATATATCTATTGGCAT . . . . GlyArgAspAsnArgThrIleIleSerLeuAsnLysTyrTyrAsn GGCAGAGATAATAGAACTATCATCAGCTTAAACAAATATTATAAT . . . . 900 LeuSerLeuHisCysLysArgProGlyAsnLysThrValLysGln CTCAGTTTGCATTGTAAGAGGCCAGGGAATAAGACAGTGAAACAA . . . . IleMetLeuMetSerGlyHisValPheHisSerHisTyrGlnPro ATAATGCTTATGTCAGGACATGTGTTTCACTCCCACTACCAGCCG . . . . . IleAsnLysArgProArgGlnAlaTrpCysTrpPheLysGluLys ATCAATAAAAGACCCAGACAAGCATGGTGCTGGTTCAAAGGCAAA 1000 . . . TrpLysAspAlaMetGlnGluValLysThrLeuAlaLysHisPro TGGAAAGACGCCATGCAGGAGGTGAAGACCCTTGCAAAACATCCC . . . . . ArgTyrArgGlyThrAsnAspThrArgAsnIleSerPheAlaAla AGGTATAGAGGAACCAATGACACAAGGAATATTAGCTTTGCAGCG . 1100 . . ProGlyLysGlySerAspProGluValAlaTyrMetTrpThrAsn CCAGGAAAAGGCTCAGACCCAGAAGTAGCATACATGTGGACTAAC . . . . . CysArgGlyGluPheLeuTyrCysAsnMetThrTrpPheLeuAsn TGCAGAGGAGAGTTTCTCTACTGCAACATGACTTGGTTCCTCAAT . . 1200 . TrpIleGluAsnLysThrHisArgAsnTyrAlaProCysHisIle TGGATAGAGAATAAGACACACCGCAATTATGCACCGTGCCATATA . . . . . LysGlnIleIleAsnThrTrpHisLysValGlyArgAsnValTyr AAGCAAATAATTAACACATGGCATAAGGTAGGGAGAAATGTATAT . . . 1300 LeuProProArgGluGlyGluLeuSerCysAsnSerThrValThr TTGCCTCCCAGGGAAGGGGAGCTGTCCTGCAACTCAACAGTAACC . . . . . SerIleIleAlaAsnIleAspTrpGlnAsnAsnAsnGlnThrAsn AGCATAATTGCTAACATTGACTGGCAAAACAATAATCAGACAAAC . . . . IleThrPheSerAlaGluValAlaGluLeuTyrArgLeuGluLeu ATTACCTTTAGTGCAGAGGTGGCAGAACTATACAGATTGGAGTTG 1400 . . . . GlyAspTyrLysLeuValGluIleThrProIleGlyPheAlaPro GGAGATTATAAATTGGTAGAAATAACACCAATTGGCTTCGCACCT . . . . ThrLysGluLysArgTyrSerSerAlaHisGlyArgHisThrArg ACAAAAGAAAAAAGATACTCCTCTGCTCACGGGAGACATACAAGA . 1500 . . . GlyValPheValLeuGlyPheLeuGlyPheLeuAlaThrAlaGly GGTGTGTTCGTGCTAGGGTTCTTGGGTTTTCTCGCAACAGCAGGT . . . . SerAlaMetGlyAlaArgAlaSerLeuThrValSerAlaGlnSer TCTGCAATGGGCGCTCGAGCGTCCCTGACCGTGTCGGCTCAGTCC . . 1600 . . ArgThrLeuLeuAlaGlyIleValGlnGlnGlnGlnGlnLeuLeu CGGACTTTACTGGCCGGGATAGTGCAGCAACAGCAACAGCTGTTG . . . . AspValValLysArgGlnGlnGluLeuLeuArgLeuThrValTrp GACGTGGTCAAGAGACAACAAGAACTGTTGCGACTGACCGTCTGG . . . 1700 . GlyThrLysAsnLeuGlnAlaArgValThrAlaIleGluLysTyr GGAACGAAAAACCTCCAGGCAAGAGTCACTGCTATAGAGAAGTAG . . . . LeuGluAspGlnAlaArgLeuAsnSerTrpGlyCysAlaPheArg CTACAGGACCAGGCGCGGCTAAATTCATGGGGATGTGCGTTTAGA . . . . 1800 GlnValCysHisThrThrValProTrpValAsnAspSerLeuAla CAAGTCTGCCACACTACTGTACCATGGGTTAATGATTCCTTAGCA . . . . ProAspTrpAspAsnMetThrTrpGlnGluTrpGluLysGlnVal CCTGACTGGGACAATATGACGTGGCAGGAATCCCAAAAACAAGTC . . . . . ArgTyrLeuGluAlaAsnIleSerLysSerLeuGluGlnAlaGln CGCTACCTGGAGGCAAATATCAGTAAAAGTTTAGAACAGGCACAA 1900 . . . IleGlnGlnGluLysAsnMetTyrGluLeuGlnLysLeuAsnSer ATTCAGCAAGAGAAAAATATGTATGAACTACAAAAATTAAATAGC . . . . . TrpAspIlePheGlyAsnTrpPheAspLeuThrSerTrpValLys TGGGATATTTTTGGCAATTGGTTTGACTTAACCTCCTGGGTCAAG . 2000 . . TyrIleGlnTyrGlyValLeuIleIleValAlaValIleALaLeu TATATTCAATATGGAGTGCTTATAATAGTAGCAGTAATAGCTTTA . . . . . ArgIleValIleTyrValValGlnMetLeuSerArgLeuArgLys AGAATAGTGATATATGTAGTACAAATGTTAAGTAGGCTTAGAAAG . . 2100 . GlyTyrArgProValPheSerSerProProGlyTyrIleGln*** GGCTATAGGCCTGTTTTCTCTTCCCCCCCCGGTTATATCCAATAG . . . . . IleHisIleHisLysAspArgGlyGlnProAlaAsnGluGluThr ATCCATATCCACAAGGACCGGGGACAGCCAGCCAACGAAGAAACA . . . 2200 GluGluAspGlyGlySerAsnGlyGlyAspArgTyrTrpProTrp CAAGAAGACGGTGGAAGCAACGGTGGAGACACATACTGGCCCTGG . . . . . ProIleAlaTyrIleHisPheLeuIleArgGlnLeuIleArgLeu GCGATAGCATATATACATTTCCTGATCCGCCAGCTGATTCGCCTC . . . . LeuThrArgLeuTyrSerIleCysArgAspLeuLeuSerArgSer TTGACCAGACTATACAGCATCTGCAGGGACTTACTATCCAGGAGC 2300 . . . . PheLeuThrLeuGlnLeuIleTyrGlnAsnLeuArgAspTrpLeu CTCCTGACCCTCCAACTCATCTACCAGAATCTCAGAGACTGGCTG . . . . ArgLeuArgThrAlaPheLeuGluTyrGlyCysGluTrpIleGln AGACTTAGAACAGCCTTCTTGCAATATGGGTGCGAGTGGATCCAA . 2400 . . . GluAlaPheGlnAlaAlaAlaArgAlaThrArgGluThrLeuAla GAAGCATTCCAGGCCGCCGCGAGGGCTACAAGAGAGACTCTTGCG . . . . GlyAlaCysArgGlyLeuTrpArgValLeuGluArgIleGlyArg GGCGCGTGCAGGGGCTTGTGGAGGGTATTGGAACGAATCGGGAGG . . 2500 . . GlyIleLeuAlaValProArgArgIleArgGlnGlyAlaGluIle CGAATACTCGCGGTTCCAAGAAGGATCAGACAGGGAGCAGAAATC . . . . AlaLeuLeu***GlyThrAlaValSerAlaGlyArgLeuTyrGlu GCCCTCCTGTGAGGGACGGCAGTATCAGCAGGGAGACTTTATGAA . . . 2600 . TyrSerMetGluGlyProSerSerArgLysGlyGluLysPheVal TACTCCATGGAAGGACCCAGCAGCAGAAAGGGAGAAAAATTTGTA . . . . GlnAlaThrLysTyrGly GAGGCAACAAAATATGGA . . - - Gag sequence - - MetGlyAlaArgAsnSerValLeuArgGlyLysLysAlaAspGlu ATGGGCGCGAGAAACTCCGTCTTGAGAGGGAAAAAAGCAGATGAA . . . . LeuGluArgIleArgLeuArgProGlyGlyLysLysLysTyrArg TTAGAAAGAATCAGGTTACGGCCCGGCGGAAAGAAAAAGTACAGG . . . . . LeuLysHisIleValTrpAlaAlaAsnLysLeuAspArgPheGly CTAAAACATATTGTGTGGGCAGCGAATAAATTGGACAGATTCGGA 100 . . . LeuAlaGluSerLeuLeuGluSerLysGluGlyCysGluLysIle TTAGCAGAGAGCCTGTTGGAGTCAAAAGAGGGTTGTCAAAAAATT . . . . . LeuThrValLeuAspProMetValProThrGlySerGluAsnLeu CTTACAGTTTTAGATCCAATGGTACCGACAGGTTCAGAAAATTTA . 200 . . LysSerLeuPheAsnThrValCysValIleTrpCysIleHisAla AAAAGTCTTTTTAATACTGTCTGCGTCATTTGGTGCATACACGCA . . . . . GluGluLysValLysAspThrGluGlyAlaLysGlnIleValArg GAAGAGAAAGTGAAAGATACTGAAGGAGCAAAACAAATAGTGCGG . . 300 . ArgHisLeuValAlaGluThrGlyThrAlaGluLysMetProSer AGACATCTAGTGGCAGAAACAGGAACTGCAGAGAAAATGCCAAGG . . . . . ThrSerArgProThrAlaProSerSerGluLysGlyGlyAsnTyr ACAAGTAGACCAACAGCACCATCTAGCGAGAAGGGAGGAAATTAC . . . 400 ProValGlnHisValGlyGlyAsnTyrThrHisIleProLeuSer CCAGTGCAACATGTAGGCGGCAACTACACCCATATACCGCTGAGT . . . . . ProArgThrLeuAsnAlaTrpValLysLeuValGluGluLysLys CCCCGAACCCTAAATGCCTGGGTAAAATTAGTAGAGGAAAAAAAG . . . . PheGlyAlaGluValValProGlyPheGluAlaLeuSerGluGly TTCGGGGCAGAAGTAGTGCCAGGATTTCAGGCACTCTCAGAAGGC 500 . . . . CysThrProTyrAspIleAsnGlnMetLeuAsnCysValGlyAsp TGGACGCCCTATGATATCAACCAAATGCTTAATTGTGTGGGCGAC . . . . HisGlnAlaAlaMetGlnIleIleArgGluIleIleAsnGluGlu CATCAAGCAGCCATGCAGATAATCAGGGAGATTATCAATGAGGAA . 600 . . . AlaAlaGluTrpAspValGlnHisProIleProGlyProLeuPro GCAGCAGAATGGGATGTGCAACATCCAATACCAGCCCCCTTAGCA . . . . AlaGlyGlnLeuArgGluProArgGlySerAspIleAlaGlyThr GCGGGGCAGCTTAGAGAGCCAAGGGGATCTGACATAGCAGGGACA . . 700 . . ThrSerThrValGluGluGlnIleGlnTrpMetPheArgProGln ACAAGCACAGTAGAAGAACAGATCCAGTGGATGTTTAGGCCACAA . . . . AsnProValProValGlyAsnIleTyrArgArgTrpIleGlnIle AATCCTGTACCAGTAGGAAACATCTATAGAAGATGGATCCAGATA . . . 800 . GlyLeuGlnLysCysValArgMetTyrAsnProThrAsnIleLeu GGATTGCAGAAGTGTGTCAGGATGTACAACCCGACCAACATCCTA . . . . AspIleLysGlnGlyProLysGluProPheGlnSerTyrValAsp GACATAAAACAGGGACCAAAGGAGCCGTTCCAAAGCTATGTAGAT . . . . 900 ArgPheTyrLysSerLeuArgAlaGluGlnThrAspProAlaVal AGATTCTACAAAAGCTTGAGGGCAGAACAAACAGATCCAGCAGTG . . . . LysAsnTrpMetThrGlnThrLeuLeuValGlnAsnAlaAsnPro AAGAATTGGATGACCCAAACACTGCTAGTACAAAATGCCAACCCA . . . . . AspCysLysLeuValLeuLysGlyLeuGlyMetAsnProThrLeu GACTGTAAATTAGTGCTAAAAGGACTAGGGATGAACCCTACCTTA 1000 . . . GluGluMetLeuThrAlaCysGlnGlyValGlyGlyProGlyGln GAAGAGATGCTGACCGCCTGTCAGGGGGTAGGTGGGCCAGGCCAG . . . . . LysAlaArgLeuMetAlaGluAlaLeuLysGluValIleGlyPro AAAGCTAGATTAATGGCAGAGGCCCTGAAAGAGGTCATAGGACCT . 1100 . . AlaProIleProPheAlaAlaAlaGlnGlnArgLysAlaPheLys GCCCCTATCCCATTCGCAGCAGCCCAGCAGAGAAAGGCATTTAAA . . . . . CysTrpAsnCysGlyLysGluGlyHisSerAlaArgGlnCysArg TGCTGGAACTGTGGAAAGGAAGGGCACTCGGCAAGACAATGCCGA . . 1200 . AlaProArgArgGlnGlyCysTrpLysCysGlyLysProGlyHis GCACCTAGAAGGCAGGGCTGCTGGAAGTGTGGTAAGCCAGGACAC . . . . . IleMetThrAsnCysProAspArgGlnAlaGlyPheLeuGlyLeu ATCATGACAAACTGCCCAGATAGACAGGCAGGTTTTTTAGGACTG . . . 1300 GlyProTrpGlyLysLysProArgAsnPheProValAlaGlnVal GGCCCTTGGGGAAAGAAGCCCCGCAACTTCCCCGTGGCCCAAGTT . . . . . ProGlnGlyLeuThrProThrAlaProProValAspProAlaVal CCGCAGGGGCTGACACCAACAGCACCCCCAGTGGATCCAGCAGTG . . . . AspLeuLeuGluLysTyrMetGlnGlnGlyLysArgGlnArgGlu GATCTACTGGAGAAATATATGCAGCAAGGGAAAAGACAGAGAGAG 1400 . . . . GlnArgGluArgProTyrLysGluValThrGluAspLeuLeuHis CAGAGAGAGAGACCATACAAGGAAGTGACAGAGGACTTACTGCAC . . . . LeuGluGlnGlyGluThrProTyrArgGlnProProThrGluAsp CTCGAGCAGGGGGAGACACCATACAGGGAGCCACCAACAGAGGAC . 1500 . . . LeuLeuHisLeuAsnSerLeuPheGlyLysAspGln TTGCTGCACCTCAATTCTCTCTTTGGAAAAGACCAG . . .
Example 6: Peptide Sequences Encoded By The ENV and GAG genes
The following coding regions for antigenic peptides, identified for convenience only by the nucleotide numbers of Example 5, within the env and gag gene regions are of particular interest.
envl (1732-1809) ArgValThrAlaIleGluLysTyr AGAGTCACTGCTATAGAGAAGTAC . . LeuGlnAspGlnAlaArgLeuAsnSerTrpGlyCysAlaPheArg CTACAGGACCAGGCGCGGCTAAATTCATGGGGATGTGCGTTTAGA . . . . 1800 GlnValCys CAAGTCTGC - env2 (1912-1983) SerLysSerLeuGluGlnAlaGln AGTAAAAGTTTAGAACAGGCACAA . . IleGlnGlnGluLysAsnMetTyrGluLeuGlnLysLeuAsnSer ATTCAGCAAGAGAAAAATATGTATGAACTACAAAAATTAAATAGC 1940 . . . . Trp TGG - env3 (1482-1530) Pro ThrLysGluLysArgTyrSerSerAlaHisGlyArgHisThrArg CCT ACAAAAGAAAAAAGATACTCCTCTGCTCACGGGAGACATACAAGA . 1500 . . . - env4 (55-129) CysThrGlnTyrValThrValPheTyrGlyValPro TGCACCCAATATGTAACTGTTTTCTATGGCGTACCC . . . . ThrTrpLysAsnAlaThrIleProLeuPheCysAlaThr ACGTGGAAAAATGCAACCATTCCCCTGTTTTGTGCAACC 100 . . - env5 (175-231) AspAsp GATGAT . TyrGlnGlnIleThrLeuAsnValThrGluAlaPheAspAlaTrp TATCAGGAAATAACTTTGAATGTAACAGAGGCTTTTGATGCATGG . 200 . . AsnAsn AATAAT - env6 (274-330) GluThrSerIleLysProCysValLysLeuThrProLeuCys GAGACATCAATAAAACCATGTGTGAAACTAACACCTTTATGT . . 300 . ValAlaMetLysCys GTAGCAATGAAATGC . . - env7 (607-660) AsnHisCysAsnThrSerValIle AACCATTGCAACACATCAGTCATC 610 . . ThrGluSerCysAspLysHisTyrTrpAsp ACAGAATCATGTGACAAGCACTATTGGGAT . . . - env8 (661-720) AlaIleArgPheArg GCTATAAGGTTTAGA . TyrCysAlaProProGlyTyrAlaLeuLeuArgCysAsnAspThr TACTGTGCACCACCGGGTTATGCCCTATTAAGATGTAATGATACC . . 700 . . - env9 (997-1044) LysArgProArgGlnAlaTrpCysTrpPheLysGlyLys AAAAGACCCAGACAAGCATGGTGCTGGTTCAAAGGCAAA 1000 . . . TrpLysAsp TGGAAAGAC - env10 (1132-1215) LysGlySerAspProGluValAlaTyrMetTrpThrAsn AAAGGCTCAGACCCAGAAGTAGCATACATGTGGACTAAC . . . . CysArgGlyGluPheLeuTyrCysAsnMetThrTrpPheLeuAsn TGCAGAGGAGAGTTTCTCTACTGCAACATGACTTGGTTCCTCAAT . . 1200 . - env11 (1237-1305) ArgAsnTyrAlaProCysHisIle CGCAATTATGCACCGTGCCATATA . . . LysGlnIleIleAsnThrTrpHisLysValGlyArgAsnValTyr AAGCAAATAATTAACACATGGCATAAGGTAGGGAGAAATGTATAT . . . 1300 - gagl (991-1053) AspCysLysLeuValLeuLysGlyLeuGlyMetAsnProThrLeu GACTGTAAATTAGTGCTAAAAGGACTAGGGATGAACCCTACCTTA 1000 . . . GluGluMetLeuThrAla GAAGAGATGCTGACCGCC . .
Of the foregoing peptides, env1, env2, env3 and gag1 are particularly contemplated for diagnostic purposes, and env4, env5, env6, env7, env8, env9, env10 and env11 are particularly contemplated as protecting agents. These peptides have been selected in part because of their sequence homology to certain of the envelope and gag protein products of other of the retroviruses in the HIV group. For vaccinating purposes, the fore going peptides may be coupled to a carrier protein by utilizing suitable and well known techniques to enhance the host's immune response. Adjuvants such as calcium phosphate or alum hydroxide may also be added. The foregoing peptides can be synthesized by conventional protein synthesis techniques, such as that of Merrifield.
It will be apparent to those skilled in the art that various modifications and variations can be made in the processes and products of the present invention. Thus, it is intended that the present application cover the modifications and variations of this invention provided they come within the scope of the appended claims and their equivalents. For convenience in interpreting the following claims, the following table sets forth the correspondence between codon codes and amino acids and the correspondence between three-letter and one-letter amino acid symbols.
__________________________________________________________________________ DNA CODON AMINO ACID 3 LET. AMINO ACID 1 LET.__________________________________________________________________________ .backslash.2 T C A G T C A G T C A G - 1 3.backslash. - T TTT TCT TAT TGT PHE SER TYR CYS F S Y C - T C TTC TCC TAC TGC PHE SER TYR CYS F S Y C - A TTA TCA TAA TGA LEU SER *** *** L S * * - G TTG TCG TAG TGG LEU SER *** TRP L S * W - T CTT CCT CAT CGT LEU PRO HIS ARG L P H R - C C CTC CCC CAC CGC LEU PRO HIS ARG L P H R - A CTA CCA CAA CGA LEU PRO GLN ARG L P Q R - G CTG CCG CAG CGG LEU PRO GLN ARG L P Q R - T ATT ACT AAT AGT ILE THR ASN SER I T N S - A C ATC ACC AAC AGC ILE THR ASN SER I T N S - A ATA ACA AAA AGA ILE THR LYS ARG I T K R - G ATG ACG AAG AGG MET THR LYS ARG M T K R - T GTT GCT GAT GGT VAL ALA ASP GLY V A D G - G C GTC GCC GAC GGC VAL ALA ASP GLY V A D G - A GTA GCA GAA GGA VAL ALA GLU GLY V A E G - G GTG GCG GAG GGG VAL ALA GLU GLY V A E G__________________________________________________________________________ -3 Letter 1 Letter CODONS__________________________________________________________________________ - ALA A GCT GCC GCA GCG - ARG R CGT CGC CGA CGG AGA AGG - ASN N AAT AAC - ASP D GAT GAC - CYS C TGT TCC - GLN Q CAA CAG - GLU E GAA GAG - GLY G GGT GGC GGA GGG - HIS H CAT CAC - ILE I ATT ATC ATA - LEU L CTT CTC CTA CTG TTA TTG - LYS K AAA AAG - MET M ATG - PHE F TTT TTC - PRO P CCT CCC CCA CCG - SER S TCT TCC TCA TCG AGT AGC - THR T ACT ACC ACA ACG - TRP H TGG - TYR Y TAT TAC - VAL V GTT GTC GTA GTG - *** * TAA TAG TGA__________________________________________________________________________
Claims
  • 1. A method for determining the effectiveness of an agent for inhibiting a Human Immunodeficiency Virus Type 2 (HIV-2) from binding to a target cell, said method comprising:
  • contacting said cell with said agent to be tested in the presence and in the absence of a labeled peptide having immunological properties of a first portion of the envelope glycoprotein of an HIV-2 virus, wherein said peptide comprises no more than about 40 amino acid residues, said first portion of the envelope glycoprotein is antigenic or is capable of eliciting the production of antibodies directed to the peptide, and said envelope glycoprotein comprises an amino acid sequence as follows:
  • MetMetAsnGlnLeuLeuIleAlaIleLeuLeuAlaSerAlaCys - 10 - LeuValTyrCysThrGlnTyrValThrValPheTyrGlyValPro - 20 30 - ThrTrpLysAsnAlaThrIleProLeuPheCysAlaThrArgAsn - 40 - ArgAspThrTrpGlyThrIleGlnCysLeuProAspAsnAspAsp - 50 60 - TryGlnGluIleThrLeuAsnValThrGluAlaPheAspAlaTrp - 70 - AsnAsnThrValThrGluGlnAlaIleGluAspValTrpHisLeu - 80 90 - PheGluThrSerIleLysProCysValLysLeuThrProLeuCys - 100 - ValAlaMetLysCysSerSerThrGluSerSerThrGlyAsnAsn - 110 120 - ThrThrSerLysSerThrSerThrThrThrThrThrProThrAsp - 130 - GlnGluGlnGluIleSerGluAspThrProCysAlaArgAlaAsp - 140 150 - AsnCysSerGlyLeuGlyGluGluGluThrIleAsnCysGlnPhe - 160 - AsnMetThrGlyLeuGluArgAspLysLysLysGlnTyrAsnGlu - 170 180 - ThrTrpTyrSerLysAspValValCysGluThrAsnAsnSerThr - 190 - AsnGlnThrGlnCysTyrMetAsnHisCysAsnThrSerValIle - 200 210 - ThrGluSerCysAspLysHisTyrTrpAspAlaIleArgPheArg - 220 - TyrCysAlaProProGlyTyrAlaLeuLeuArgCysAsnAspThr - 230 240 - AsnTyrSerGlyPheAlaProAsnCysSerLysValValAlaSer - 250 - ThrCysThrArgMetMetGluThrGlnThrSerThrTrpPheGly - 260 270 - PheAsnGlyThrArgAlaGluAsnArgThrTyrIleTyrTrpHis - 280 - GlyArgAspAsnArgThrIleIleSerLeuAsnLysTryTyrAsn - 290 300 - LeuSerLeuHisCysLysArgProGlyAsnLysThrValLysGln - 310 - IleMetLeuMetSerGlyHisValPheHisSerHisTyrGlnPro - 320 340 - IleAsnLysArgProArgGlnAlaTrpCysTrpPheLysGlyLys - 350 - TrpLysAspAlaMetGlnGluValLysThrLeuAlaLysHisPro - 360 370 - ArgTyrArgGlyThrAsnAspThrArgAsnIleSerPheAlaAla - 380 - ProGlyLysGlySerAspProGluValAlaTyrMetTrpThrAsn - 390 400 - CysArgGlyGluPheLeuTyrCysAsnMetThrTrpPheLeuAsn - 410 - TrpIleGluAsnLysThrHisArgAsnTyrAlaProCysHisIle - 420 430 - LysGlnIleIleAsnThrTrpHisLysValGlyArgAsnValTyr - 440 - LeuProProArgGluGlyGluLeuSerCysAsnSerThrValThr - 450 460 - SerIleIleAlaAsnIleAspTrpGlnAsnAsnAsnGlnThrAsn - 470 - IleThrPheSerAlaGluValAlaGluLeuTyrArgLeuGluLeu - 480 490 - GlyAspTyrLysLeuValGluIleThrProIleGlyPheAlaPro - 500 - ThrLysGluLysArgTyrSerSerAlaHisGlyArgHisThrArg - 510 520 - GlyValPheValLeuGlyPheLeuGlyPheLeuAlaThrAlaGly - 530 - SerAlaMetGlyAlaArgAlaSerLeuThrValSerAlaGlnSer - 540 550 - ArgThrLeuLeuAlaGlyIleValGlnGlnGlnGlnGlnLeuLeu - 560 - AspValValLysArgGlnGlnGluLeuLeuArgLeuThrValTrp - 570 580 - GlyThrLysAsnLeuGlnAlaArgValThrAlaIleGluLysTyr - 590 - LeuGluAspGlnAlaArgLeuAsnSerTrpGlyCysAlaPheArg - 600 610 - GlnValCysHisThrThrValProTrpValAsnAspSerLeuAla - 620 - ProAspTrpAspAsnMetThrTrpGlnGluTrpGluLysGlnVal - 630 640 - ArgTyrLeuGluAlaAsnIleSerLysSerLeuGluGluAlaGln - 650 - IleGlnGlnGluLysAsnMetTyrGluLeuGlnLysLeuAsnSer - 660 670 - TrpAspIlePheGlyAsnTrpPheAspLeuThrSerTrpValLys - 680 - TyrIleGlnTyrGlyValLeuIleIleValAlaValIleAlaLeu - 690 700 - ArgIleValIleTyrValValGlnMetLeuSerArgLeuArgLys - 710 - GlyTyrArgProValPheSerSerProProGlyTyrIleGln*** - 720 730 - IleHisIleHisLysAspArgGlyGlnProAlaAsnGluGluThr - 740 - GluGluAspGlyGlySerAsnGlyGlyAspArgTyrTrpProTrp - 750 760 - ProIleAlaTyrIleHisPheLeuIleArgGlnLeuIleArgLeu - 770 - LeuThrArgLeuTyrSerIleCysArgAspLeuLeuSerArgSer - 780 790 - PheLeuThrLeuGlnLeuIleTyrGlnAsnLeuArgAspTrpLeu - 800 - ArgLeuArgThrAlaPheLeuGlnTyrGlyCysGluTrpIleGln - 810 820 - GluAlaPheGlnAlaAlaAlaArgAlaThrArgGluThrLeuAla - 830 - GlyAlaCysArgGlyLeuTrpArgValLeuGluArgIleGlyArg - 840 850 - GlyIleLeuAlaValProArgArgIleArgGlnGlyAlaGluIle - 850 - AlaLeuLeu***GlyThrAlaValSerAlaGlyArgLeuTyrGlu - 860 870 - TyrSerMetGluGlyProSerSerArgLysGlyGluLysPheVal - 880 - GlnAlaThrLysTyrGly; - 890
  • determining the amount of labeled peptide bound in the presence of said agent;
  • determining the amount of labeled peptide bound in the absence of said agent; and
  • determining the relative amount of labeled peptide-cell binding in the presence of said agent compared to the amount of labeled peptide-cell binding in the absence of said agent;
  • wherein an agent having a low affinity for binding to the cell is evidenced by little or no change between the binding values of the labeled peptide in the presence and in the absence of the agent, and an agent having a high affinity for binding to the cell is evidenced by a lower binding value in the presence of the agent in comparison to the binding value in the absence of the agent.
  • 2. The method of claim 1, wherein said enveloped glycoprotein is labeled with an immunoassay label selected from the group consisting of enzymes, radioactive isotopes, fluorescent labels, and chromophores.
  • 3. A method for determining the effectiveness of an agent for inhibiting a Human Immunodeficiency Virus Type 2 (HIV-2) from binding to a target cell, said method comprising:
  • contacting said cell with said agent to be tested in the presence of in the absence of one or more labeled peptides selected from the group consisting of:
  • (1) a peptide comprising an amino acid sequence of either of the following formulas:
  • XR--A-E-YL-DQ--L--WGC-----CZ, or
  • XA-E-YL-DZ,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of either of the following peptide sequences:
  • RVTAIEKYLQDQARLNSWGCAFRQVC, or
  • AIEKYLQDQ;
  • (2) a peptide comprising an amino acid sequence of either of the following formulas:
  • X----E--Q-QQEKN--EL--L---Z, or
  • XQ-QQEKNZ,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of either of the following peptide sequences:
  • SKSLEQAQIQQEKNMYELQKLNSW, or
  • QIQQEKN;
  • (3) a peptide comprising an amino acid sequence of either of the following formulas:
  • XEL--YK-V-I-P-G-APTK-KR-----Z, or
  • XYK-V-I-P-G-APTK-KRZ,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of either of the following peptide sequences:
  • ELGDYKLVEITPIGFAPTKEKRYSSAH, or
  • YKLVEITPIGFAPTKEK;
  • (4) the antigenic peptide gagl comprising an amino acid sequence of the following formula:
  • XNCKLVLKGLGMNPTLEEMLTAZ,
  • wherein X and Z or OH or NH2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of the following peptide sequence:
  • XNCKLVLKGLGMNPTLEEMLTA;
  • (5) a peptide comprising an amino acid sequence of either of the following formulas:
  • X----VTV-YGVP-WK-AT--LFCA-Z, or
  • XVTV-YGVP-WK-ATZ,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of either of the following peptide sequences:
  • CTQYVTVFYGVPTWKNATIPLFCAT, or VTVFYGVPTWKNAT;
  • (6) a peptide comprising an amino acid sequence of either of the following formulas:
  • X-G-DPE------NC-GEF-YCN-----NZ, or
  • XNC-GEF-YCNZ,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of either of the following peptide sequences:
  • KGSDPEVAYMWTNCRGEFLYCNMTWFLN, or
  • NCRGEFLYCN;
  • (7) a peptide comprising an amino acid sequence of either of the following formulas:
  • X-----C-IKQ-I------G---YZ, or
  • XC-IKQ-IZ,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of either of the following peptide sequences:
  • RNYAPCHIKQIINTWHKVGRNVY, or
  • CHIKQII;
  • (8) a peptide comprising an amino acid sequence of either of the following formulas:
  • X---QE--LNVTE-F--W-NZ, or
  • XLNVTE-FZ,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of the following peptide sequence:
  • DDYQEITLNVTEAFDAWNN;
  • (9) a peptide comprising an amino acid sequence of either of the following formulas:
  • XL---S-KPCVKLTPLCV--KZ, or
  • XKPCVKLTPLCVZ, or
  • XS-KPCVKLTPLCVZ,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of the following peptide sequence:
  • ETSIKPCVKLTPLCVAMK;
  • (10) a peptide comprising an amino acid sequence of either of the following formulas:
  • X---N-S-IT--C-Z, or
  • XN-S-ITZ,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of the following peptide sequence:
  • NHCNTSVITESCD;
  • (11) a peptide comprising an amino acid sequence having the following formula:
  • XYC-P-G-A-L-CN-TZ,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of the following peptide sequence:
  • YCAPPGYALLRCNDT; and
  • (12) a peptide comprising an amino acid sequence of either of the following formulas:
  • X------A-C-----W--Z,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of the following peptide sequence:
  • NKRPRQAWCWFKGKWKD;
  • wherein said immunological properties comprise the ability of said peptide sequences to specifically recognize antibodies against HIV-2;
  • determining the amount of labeled peptide bound in the presence of said agent;
  • determining the amount of labeled peptide bound in the absence of said agent; and
  • determining the relative amount of labeled peptide-cell binding in the presence of said agent compared to the amount of labeled peptide-cell binding in the absence of said agent;
  • wherein an agent having a low affinity for binding to the cell is evidenced by little or no change between the binding values of the labeled peptide in the presence and in the absence of the agent, and an agent having a high affinity for binding to the cell is evidenced by a lower binding value in the presence of the agent in comparison to the binding value in the agent in comparison to the binding value in the absence of the agent.
  • 4. The method of claim 3, wherein said one or more peptides are labeled with an immunoassay label selected from the group consisting of enzymes, radioactive isotopes, fluorescent labels, and chromophores.
  • 5. A kit for determining the effectiveness of an agent for inhibiting a Human Immunodeficiency Virus Type 2 (HIV-2) from binding to a target cell, said kit comprising:
  • a labeled peptide having immunological properties of a first portion of the envelope glycoprotein of a HIV-2 virus, wherein said peptide comprises no more than about 40 amino acid residues, said first portion of the envelope glycoprotein is antigenic or is capable of eliciting the production of antibodies directed to the peptide, and said envelope glycoprotein comprises an amino acid sequence as follows:
  • MetMetAsnGlnLeuLeuIleAlaIleLeuLeuAlaSerAlaCys 10 - LeuValTyrCysThrGlnTyrValThrValPheTyrGlyValPro 20 30 - ThrTrpLysAsnAlaThrIleProLeuPheCysAlaThrArgAsn 40 - ArgAspThrTrpGlyThrIleGlnCysLeuProAspAsnAspAsp 50 60 - TryGlnGluIleThrLeuAsnValThrGluAlaPheAspAlaTrp 70 - AsnAsnThrValThrGluGlnAlaIleGluAspValTrpHisLeu 80 90 - PheGluThrSerIleLysProCysValLysLeuThrProLeuCys 100 - ValAlaMetLysCysSerSerThrGluSerSerThrGlyAsnAsn 110 120 - ThrThrSerLysSerThrSerThrThrThrThrThrProThrAsp 130 - GlnGluGlnGluIleSerGluAspThrProCysAlaArgAlaAsp 140 150 - AsnCysSerGlyLeuGlyGluGluGluThrIleAsnCysGlnPhe 160 - AsnMetThrGlyLeuGluArgAspLysLysLysGlnTyrAsnGlu 170 180 - ThrTrpTyrSerLysAspValValCysGluThrAsnAsnSerThr 190 - AsnGlnThrGlnCysTyrMetAsnHisCysAsnThrSerValIle 200 210 - ThrGluSerCysAspLysHisTyrTrpAspAlaIleArgPheArg 220 - TyrCysAlaProProGlyTyrAlaLeuLeuArgCysAsnAspThr 230 240 - AsnTyrSerGlyPheAlaProAsnCysSerLysValValAlaSer 250 - ThrCysThrArgMetMetGluThrGlnThrSerThrTrpPheGly 260 270 - PheAsnGlyThrArgAlaGluAsnArgThrTyrIleTyrTrpHis 280 - GlyArgAspAsnArgThrIleIleSerLeuAsnLysTryTyrAsn 290 300 - LeuSerLeuHisCysLysArgProGlyAsnLysThrValLysGln 310 - IleMetLeuMetSerGlyHisValPheHisSerHisTyrGlnPro 320 340 - IleAsnLysArgProArgGlnAlaTrpCysTrpPheLysGlyLys 350 - TrpLysAspAlaMetGlnGluValLysThrLeuAlaLysHisPro 360 370 - ArgTyrArgGlyThrAsnAspThrArgAsnIleSerPheAlaAla 380 - ProGlyLysGlySerAspProGluValAlaTyrMetTrpThrAsn 390 400 - CysArgGlyGluPheLeuTyrCysAsnMetThrTrpPheLeuAsn 410 - TrpIleGluAsnLysThrHisArgAsnTyrAlaProCysHisIle 420 430 - LysGlnIleIleAsnThrTrpHisLysValGlyArgAsnValTyr 440 - LeuProProArgGluGlyGluLeuSerCysAsnSerThrValThr 450 460 - SerIleIleAlaAsnIleAspTrpGlnAsnAsnAsnGlnThrAsn 470 - IleThrPheSerAlaGluValAlaGluLeuTyrArgLeuGluLeu 480 490 - GlyAspTyrLysLeuValGluIleThrProIleGlyPheAlaPro 500 - ThrLysGluLysArgTyrSerSerAlaHisGlyArgHisThrArg 510 520 - GlyValPheValLeuGlyPheLeuGlyPheLeuAlaThrAlaGly 530 - SerAlaMetGlyAlaArgAlaSerLeuThrValSerAlaGlnSer 540 550 - ArgThrLeuLeuAlaGlyIleValGlnGlnGlnGlnGlnLeuLeu 560 - AspValValLysArgGlnGlnGluLeuLeuArgLeuThrValTrp 570 580 - GlyThrLysAsnLeuGlnAlaArgValThrAlaIleGluLysTyr 590 - LeuGluAspGlnAlaArgLeuAsnSerTrpGlyCysAlaPheArg 600 610 - GlnValCysHisThrThrValProTrpValAsnAspSerLeuAla 620 - ProAspTrpAspAsnMetThrTrpGlnGluTrpGluLysGlnVal 630 640 - ArgTyrLeuGluAlaAsnIleSerLysSerLeuGluGluAlaGln 650 - IleGlnGlnGluLysAsnMetTyrGluLeuGlnLysLeuAsnSer 660 670 - TrpAspIlePheGlyAsnTrpPheAspLeuThrSerTrpValLys 680 - TyrIleGlnTyrGlyValLeuIleIleValAlaValIleAlaLeu 690 700 - ArgIleValIleTyrValValGlnMetLeuSerArgLeuArgLys 710 - GlyTyrArgProValPheSerSerProProGlyTyrIleGln*** 720 730 - IleHisIleHisLysAspArgGlyGlnProAlaAsnGluGluThr 740 - GluGluAspGlyGlySerAsnGlyGlyAspArgTyrTrpProTrp 750 760 - ProIleAlaTyrIleHisPheLeuIleArgGlnLeuIleArgLeu 770 - LeuThrArgLeuTyrSerIleCysArgAspLeuLeuSerArgSer 780 790 - PheLeuThrLeuGlnLeuIleTyrGlnAsnLeuArgAspTrpLeu 800 - ArgLeuArgThrAlaPheLeuGlnTyrGlyCysGluTrpIleGln 810 820 - GluAlaPheGlnAlaAlaAlaArgAlaThrArgGluThrLeuAla 830 - GlyAlaCysArgGlyLeuTrpArgValLeuGluArgIleGlyArg 840 850 - GlyIleLeuAlaValProArgArgIleArgGlnGlyAlaGluIle 850 - AlaLeuLeu***GlyThrAlaValSerAlaGlyArgLeuTyrGlu 860 870 - TyrSerMetGluGlyProSerSerArgLysGlyGluLysPheVal 880 - GlnAlaThrLysTyrGly; 890
  • a composition comprising a target cell; and
  • a control composition comprising said labeled peptide and said cell;
  • wherein the relative affinity of the agent for binding to the cell is determined by comparing the amount of labeled peptide bound in the presence of the agent with the amount of labeled peptide bound in the absence of said agent.
  • 6. The kit of claim 5, wherein said enveloper glycoprotein is labeled with an immunoassay label selected from the group consisting of enzymes, radioactive isotopes, fluorescent labels, and chromophores.
  • 7. A kit for determining the effectiveness of an agent for inhibiting a Human Immunodeficiency Virus Type 2 (HIV-2) from binding to a target cell, said kit comprising:
  • one or more labeled peptides selected from the group consisting of:
  • (1) a peptide comprising an amino acid sequence of either of the following formulas:
  • XR--A-E-YL-DQ--L--WGC-----CZ, or
  • XA-E-YL-DZ,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of either of the following peptide sequences:
  • RVTAIEKYLQDQARLNSWGCAFRQVC, or
  • AIEKYLQDQ;
  • (2) a peptide comprising an amino acid sequence of either of the following formulas:
  • X ----E--Q-QQEKN--EL--L---Z, or
  • XQ-QQEKNZ,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of either of the following peptide sequences:
  • SKSLEQAQIQQEKNMYELQKLNSW, or
  • QIQQEKN;
  • (3) a peptide comprising an amino acid sequence of either of the following formulas:
  • XEL--YK-V-I-P-G-APTK-KR-----Z, or
  • XYK-V-I-P-G-APTK-KRZ,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of either of the following peptide sequences:
  • ELGDYKLVEITPIGFAPTKEKRYSSAH, or
  • YKLVEITPIGFAPTKEK;
  • (4) the antigenic peptide gagl comprising an amino acid sequence of the following formula:
  • XNCKLVLKGLGMNPTLEEMLTAZ,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of the following peptide sequence:
  • XNCKLVLKGLGMNPTLEEMLTA;
  • (5) a peptide comprising an amino acid sequence of either of the following formulas:
  • X----VTV-YGVP-WK-AT--LFCA-Z, or
  • XVTV-YGVP-WK-ATZ,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of either of the following peptide sequences:
  • CTQYVTVFYGVPTWKNATIPLFCAT, or
  • VTVFYGVPTWKNAT;
  • (6) a peptide comprising an amino acid sequence of either of the following formulas:
  • X-G-DPE------NC-GEF-YCN-----NZ, or
  • XNC-GEF-YCNZ,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of either of the following peptide sequences:
  • KGSDPEVAYMWTNCRGEFLYCNMTWFLN, or
  • NCRGEFLYCN;
  • (7) a peptide comprising an amino acid sequence of either of the following formulas:
  • X-----C-IKQ-I------G---YZ, or
  • XC-IKQ-IZ,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of either of the following peptide sequences:
  • RNYAPCHIKQIINTWHKVGRNVY, or
  • CHIKQII;
  • (8) a peptide comprising an amino acid sequence of either of the following formulas:
  • X---QE--LNVTE-F--W-NZ, or
  • XLNVTE-FZ,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of the following peptide sequence:
  • DDYQEITLNVTEAFDAWNN;
  • (9) a peptide comprising an amino acid sequence of either of the following formulas:
  • XL---S-KPCVKLTPLCV--KZ, or
  • XKPCVKLTPLCVZ, or
  • XS-KPCVKLTPLCVZ,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of the following peptide sequence:
  • ETSIKPCVKLTPLCVAMK;
  • (10) a peptide comprising an amino acid sequence of either of the following formulas:
  • X---N-S-IT--C-Z, or
  • XN-S-ITZ,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of the following peptide sequence:
  • NHCNTSVITESCD;
  • (11) a peptide comprising an amino acid sequence having the following formula:
  • XYC-P-G-A-L-CN-TZ,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of the following peptide sequence:
  • YCAPPGYALLRCNDT; and
  • (12) a peptide comprising an amino acid sequence of either of the following formulas:
  • X------A-C-----W--Z,
  • wherein X and Z are OH or NH.sub.2, and wherein each of the hyphens corresponds to an aminoacyl residue selected from the group consisting of those which permit the conservation of the immunological properties of the following peptide sequence:
  • NKRPRQAWCWFKGKWKD;
  • wherein said immunological properties comprise the ability of said peptide sequences to specifically recognize antibodies against HIV-2;
  • a composition comprising a target cell; and
  • a control composition comprising said one or more labeled peptides and said cell;
  • wherein the relative affinity of the agent for binding to the cell is determined by comparing the amount of labeled peptide bound in the presence of the agent with the amount of labeled peptide bound in the absence of said agent.
  • 8. The kit of claim 7, wherein said one or more peptide are labeled with an immunoassay label selected from the group consisting of enzymes, radioactive isotopes, fluorescent labels, and chromophores.
  • 9. A method for determining the effectiveness of an agent for inhibiting a Human Immunodeficiency Virus Type 2 (HIV-2) from binding to a target cell, said method comprising:
  • contacting said cell with said agent to be tested in the presence and in the absence of one or more labeled polypeptide expression products of HIV-2 selected from the group consisting of p16, p26, p12, polymerase, Q protein, R protein, X protein, env protein, F protein, TAT, ART, U5, and U3;
  • determining the amount of labeled polypeptide expression product bound in the presence of said agent;
  • determining the amount of labeled polypeptide expression product bound in the absence of said agent; and
  • determining the relative amount of labeled polypeptide expression product-cell binding in the presence of said agent compared to the amount of labeled polypeptide expression product-cell binding in the absence of said agent;
  • wherein an agent having a low affinity for binding to the cell is evidenced by little or no change between the binding values of the labeled polypeptide expression product in the presence and in the absence of the agent, and an agent having a high affinity for binding to the cell is evidenced by a lower binding value in the presence of the agent in comparison to the binding value in the absence of the agent.
  • 10. The method of claim 9, wherein said env protein is labeled with an immunoassay label selected from the group consisting of enzymes, radioactive isotopes, fluorescent labels, and chromophores.
  • 11. A kit for determining the effectiveness of an agent for inhibiting a Human Immunodeficiency Virus Type 2 (HIV-2) from binding to a target cell, said kit comprising:
  • one or more labeled polypeptide expression products of HIV-2 selected from the group consisting of p16, p26, p12, polymerase, Q protein, R protein, X protein, env protein, F protein, TAT, ART, U5, and U3;
  • a composition comprising a target cell; and
  • a control composition comprising said one or more labeled polypeptide expression products and said cell;
  • wherein the relative affinity of the agent for binding to the cell is determined by comparing the amount of labeled polypeptide expression product bound in the presence of the agent with the amount of labeled polypeptide expression product bound in the absence of said agent.
  • 12. The kit of claim 11, wherein said env protein is labeled with an immunoassay label selected from the group consisting of enzymes, radioactive isotopes, fluorescent labels, and chromophores.
Priority Claims (5)
Number Date Country Kind
86 00911 Jan 1986 FRX
86 01635 Feb 1986 FRX
86 01985 Feb 1986 FRX
86 03881 Mar 1986 FRX
86 04215 Mar 1986 FRX
BACKGROUND OF THE INVENTION

This application is a division of application Ser. No. 07/752,368, filed Sep. 3, 1991, which is a division of application Ser. No. 013,477, filed Feb. 11, 1987, now U.S. Pat. No. 5,079,342, issued Jan. 7, 1992, which is continuation-in-part of allowed U.S. patent application Ser. No. 07/003,764 of Alizon et al. for "Cloned DNA Sequences Related to the Entire Genomic RNA of Human Immunodeficiency Virus II (HIV-2), Polypeptides Encoded by these DNA Sequences and Use of these DNA Clones and Polypeptides in Diagnostic Kits," filed Jan. 16, 1987, now U.S. Pat. No. 5,051,496, which is a continuation-in-part of U.S. patent application Ser. No. 06/933,184 filed Nov. 21, 1986, now abandoned in favor of continuation application Ser. No. 604,323, filed Oct. 24, 1990, now abandoned in favor of continuation application Ser. No. 732,748, filed Jul. 18, 1991 (pending) which is a continuation-in-part application of U.S. patent application Ser. No. 916,080 of Montagnier et al. for "Cloned DNA Sequences Related to the Genomic RNA of the Human Immunodeficiency Virus II (HIV-2), Poly-peptides Encoded by these DNA Sequences and Use of these DNA Clones and Polypeptides in Diagnostic Kits," filed Oct. 6, 1986, now abandoned in favor of continuation application Ser. No. 602,383, filed Oct. 24, 1990 (now abandoned), and U.S. patent application Ser. No. 835,228 of Montagnier et al. for "New Retrovirus Capable of Causing AIDS, Antigens Obtained from this Retrovirus and Corresponding Antibodies and their Application for Diagnostic Purposes," filed Mar. 3, 1986 (now U.S. Pat. No. 4,839,288, issued Jun. 13, 1989). The disclosures of each of these predecessor applications are expressly -incorporated herein by reference.

US Referenced Citations (3)
Number Name Date Kind
4629783 Cosand Dec 1986
4839288 Montagnier et al. Jun 1989
5079342 Alizon et al. Jan 1992
Foreign Referenced Citations (2)
Number Date Country
0 316 695 B1 Mar 1993 EPX
WO 8504897 Nov 1985 WOX
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Entry
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Allan et al., "Major Glycoprotein Antigens That Induce Antibodies in AIDS Patients Are Encoded by HTLV-III", Science 228, pp. 1091-1094 (1985).
Chang et al., "Detection of Antibodies to Human T-Cell Lymphotropic Virus-III (HTLV-III)with an Immunoassay Employing a Recombinant Escherichia coli-Derived Viral Antigenic Peptide", Bio/Technology, 3, pp. 905-909 (1985).
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Related Publications (1)
Number Date Country
835228 Mar 1986
Divisions (2)
Number Date Country
Parent 752368 Sep 1991
Parent 013477 Feb 1987
Continuation in Parts (3)
Number Date Country
Parent 003764 Jan 1987
Parent 933184 Nov 1986
Parent 916080 Oct 1986