Claims
- 1. A method for identifying an immunogenic peptide representative of a structural element of a target protein having a known three dimensional structure, comprising:
selecting epitopes of the target protein based on the three dimensional structure of the target protein, thereby obtaining selected epitopes; detecting, in a molecular model of a polypeptide comprising a selected epitope linked to a scaffold protein, an epitope having a three dimensional conformation corresponding to the three dimensional structure of the epitope in the target protein, thereby identifying a candidate immunogenic peptide representative of a structural element of the target protein; and detecting that antibodies induced by the candidate immunogenic peptide selectively bind to the target protein, thereby identifying an immunogenic peptide representative of a structural element of the target protein.
- 2. The method of claim 1, wherein the scaffold protein comprises a viral coat protein.
- 3. The method of claim 2, wherein the viral coat protein comprises a Hepatitis B core protein.
- 4. The method of claim 1, wherein the selected epitope is linked to the scaffold protein via a linker moiety.
- 5. The method of claim 4, wherein the linker moiety comprises a peptide.
- 6. The method of claim 1, wherein the polypeptide comprising a selected epitope linked to a scaffold protein is a fusion protein.
- 7. The method of claim 1, wherein the scaffold protein comprises a poly(amino acid).
- 8. The method of claim 7, wherein the poly(amino acid) comprises poly(lysine).
- 9. The method of claim 1, wherein the target protein comprises a protein of an infectious microorganism.
- 10. The method of claim 9, wherein the infectious microorganism is a bacterium.
- 11. The method of claim 10, wherein the infectious microorganism causes anthrax.
- 12. The method of claim 11, wherein the target protein comprises anthrax protective antigen.
- 13. The method of claim 11, wherein the target protein comprises anthrax lethal factor.
- 14. The method of claim 2, further comprising assembling a plurality of immunogenic peptides, each immunogenic peptide comprising an epitope linked to a viral coat protein, into a virus-like particle.
- 15. The method of claim 14, wherein epitopes of the immunogenic peptides of the plurality are the same.
- 16. The method of claim 14, wherein epitopes of the immunogenic peptides of the plurality comprise anthrax protective antigen epitopes and anthrax lethal factor epitopes.
- 17. An immunogenic peptide identified according to the method of claim 1.
- 18. An isolated peptide, consisting of:
amino acid residues 606 to 705 of SEQ ID NO:30; amino acid residues 606 to 735 of SEQ ID NO:30; amino acid residues 606 to 706 of SEQ ID NO:30; amino acid residues 606 to 704 of SEQ ID NO:30; amino acid residues 606 to 734 of SEQ ID NO:30; amino acid residues 607 to 703 of SEQ ID NO:30; amino acid residues 606 to 732 of SEQ ID NO:30; amino acid residues 604 to 707 of SEQ ID NO:30; amino acid residues 606 to 730 of SEQ ID NO:30; or amino acid residues 606 to 733 of SEQ ID NO:30.
- 19. An isolated antibody that selectively binds a peptide of claim 18, provided said antibody does not substantially bind a peptide comprising:
amino acid residues 596 to 735 of SEQ ID NO:30; amino acid residues 679 to 693 of SEQ ID NO:30; amino acid residues 703 to 722 of SEQ ID NO:30; or amino acid residues 671 to 721 of SEQ ID NO:30.
- 20. An isolated polynucleotide encoding the peptide of claim 18.
- 21. An isolated peptide, consisting of:
amino acid residues 17 to 153 of SEQ ID NO:30; amino acid residues 261 to 454 of SEQ ID NO:30; or amino acid residues 487 to 594 of SEQ ID NO:30.
- 22. A composition, comprising at least a first peptide of claim 18 operatively linked to at least a first heterologous molecule.
- 23. The composition of claim 22, wherein the first heterologous molecule comprises a linker moiety.
- 24. The composition of claim 23, wherein the linker moiety comprises a peptide linker.
- 25. The composition of claim 24, wherein the peptide linker comprises an oligo(glycine) linker.
- 26. The composition of claim 24, wherein the peptide linker comprises a glycine-serine-alanine linker.
- 27. The composition of claim 22, wherein the first heterologous molecule comprises a heterologous peptide.
- 28. The composition of claim 27, which comprises a fusion protein.
- 29. The composition of claim 27, wherein the heterologous peptide comprises a scaffold protein.
- 30. The composition of claim 29, wherein the scaffold protein comprises a viral coat protein.
- 31. The composition of claim 30, wherein the viral coat protein comprises a Hepatitis B core protein.
- 32. The composition of claim 31, wherein the Hepatitis B core protein consists of amino acid residues 1 to 77 and amino acid residues 80 to 149 of SEQ ID NO:34, and wherein the peptide of claim B 1 is operatively linked to amino acid residue 77 and to amino acid residue 80 of SEQ ID NO:34.
- 33. The composition of claim 32, wherein the peptide of claim 18 is operatively linked to amino acid residue 77 via a first linker moiety and is operatively linked to amino acid residue 80 via a second linker moiety.
- 34. The method of claim 33, wherein the first linker moiety and the second linker moiety are the same.
- 35. The method of claim 33, wherein each of the first linker moiety and the second linker moiety is a peptide.
- 36. The composition of claim 22, further comprising at least a second heterologous molecule.
- 37. The composition of claim 36, wherein at least the first heterologous molecule comprises a linker moiety or a scaffold protein.
- 38. The composition of claim 37, wherein the first heterologous molecule comprises a linker moiety and the second heterologous molecule comprises a scaffold protein, and wherein the peptide of claim 18 is operatively linked to the scaffold protein via the linker moiety.
- 39. The composition of claim 36, comprising, in operative linkage, a first portion of a scaffold protein, a first peptide linker, the peptide of claim 18, a second peptide linker, and a second portion of the scaffold protein.
- 40. The composition of claim 39, wherein the scaffold protein comprises a viral coat protein.
- 41. The composition of claim 39, which comprises an amino acid sequence as set forth in SEQ ID NO:36.
- 42. The composition of claim 40, which comprises a plurality of polypeptides, wherein polypeptides of the plurality comprise a peptide of claim B 1 operatively linked to a viral coat protein.
- 43. The composition of claim 42, wherein polypeptides of the plurality are assembled to form a virus-like particle.
- 44. The composition of claim 22, wherein the first heterologous molecule comprises a tag.
- 45. The composition of claim 44, wherein the tag comprises a peptide tag.
- 46. The composition of claim 45, wherein the peptide tag comprises an oligo(histidine) tag.
- 47. The composition of claim 22, which is comprises immunogenic peptide representative of a structural element of the target protein.
- 48. The composition of claim 47, further comprising a carrier.
- 49. The composition of claim 48, wherein carrier comprises an adjuvant.
- 50. A polynucleotide encoding the composition of claim 22.
- 51. The polynucleotide of claim 50, which comprises SEQ ID NO:35.
- 52. The polynucleotide of claim 50, which is contained in a vector.
- 53. The polynucleotide of claim 52, wherein the vector is a viral vector.
- 54. The polynucleotide of claim 50, which is contained in a matrix.
- 55. The polynucleotide of claim 54, wherein the matrix comprises liposomes or microbubbles.
- 56. A method of stimulating an immune response in a subject, comprising administering the composition of claim 22, or a polynucleotide encoding the composition, to a subject under conditions suitable for stimulating an immune response.
- 57. The method of claim 56, wherein the immune response comprises a protective immune response.
- 58. The method of claim 56, wherein the subject is a mammalian subject.
- 59. The method of claim 56, wherein the subject is a human subject.
- 60. A method for identifying an immunogenic peptide representative of a structural element of a target protein having a known amino acid sequence but an unknown three dimensional structure, comprising:
generating a molecular model of a three dimensional structure of the target protein based on the three dimensional structure of a homologous protein; selecting epitopes of the target protein based on the molecular model of the three dimensional structure of the target protein, thereby obtaining selected epitopes; detecting, in a molecular model of a chimeric polypeptide comprising at least one selected epitope having a constrained structure, an epitope having a three dimensional conformation corresponding to the molecular model of the three dimensional structure of the epitope in the target protein, thereby identifying a candidate immunogenic peptide representative of a structural element of the target protein; and detecting that antibodies induced by the candidate immunogenic peptide selectively bind to the target protein, thereby identifying an immunogenic peptide representative of a structural element of the target protein.
- 61. The method of claim 60, wherein the chimeric polypeptide comprises a plurality of linked selected epitopes.
- 62. The method of claim 61, wherein the selected epitopes of the plurality are the same.
- 63. The method of claim 60, wherein the chimeric polypeptide further comprises at least one linker peptide, which is linked to at least one selected epitope of the chimeric polypeptide.
- 64. The method of claim 60, wherein the constrained structure of the selected epitope is due to at least one disulfide bond in the chimeric polypeptide.
- 65. The method of claim 60, wherein the constrained structure of the selected epitope is due to cyclization of the chimeric polypeptide.
- 66. The method of claim 60, wherein the chimeric polypeptide further comprises a scaffold protein.
- 67. The method of claim 66, wherein the scaffold protein comprises a viral coat protein.
- 68. The method of claim 67, wherein the viral coat protein comprises a Hepatitis B core protein.
- 69. The method of claim 66, wherein the scaffold protein comprises a poly(amino acid).
- 70. The method of claim 69, wherein the poly(amino acid) comprises poly(lysine).
- 71. The method of claim 66, wherein the selected epitope is linked to the scaffold protein via a linker moiety.
- 72. The method of claim 71, wherein the linker moiety comprises a peptide.
- 73. The method of claim 60, wherein the target protein comprises a protein of an infectious microorganism.
- 74. The method of claim 73, wherein the infectious microorganism is a bacterium.
- 75. The method of claim 74, wherein the target protein comprises an autotransporter.
- 76. The method of claim 75, wherein the autotransporter comprises an autotransporter as set forth in Table 1.
- 77. The method of claim 74, wherein the infectious microorganism is a Shigella species.
- 78. The method of claim 77, wherein the Shigella species is Shigella flexneri.
- 79. The method of claim 77, wherein the target protein comprises a Shigella protease involved in colonization (pic) gene product.
- 80. The method of claim 74, wherein the infectious microorganism is a Yersinia species.
- 81. The method of claim 80, wherein the Yersinia species is Yersinia pestis.
- 82. The method of claim 80, wherein the target protein comprises a Yersinia murine toxin (YMt).
- 83. The method of claim 67, further comprising assembling a plurality of immunogenic peptides, each immunogenic peptide comprising an epitope linked to a viral coat protein, into a virus-like particle.
- 84. The method of claim 83, wherein epitopes of the immunogenic peptides of the plurality are the same.
- 85. An immunogenic peptide identified according to the method of claim 60.
- 86. An isolated peptide, consisting of a peptide as set forth in SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, SEQ ID NO:1 1, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:27, or SEQ ID NO:28.
- 87. An isolated polynucleotide encoding the peptide of claim 86.
- 88. The polynucleotide of claim 87, which is contained in a vector.
- 89. The polynucleotide of claim 88, wherein the vector is a viral vector.
- 90. The polynucleotide of claim 87, which is contained in a matrix.
- 91. The polynucleotide of claim 90, wherein the matrix comprises liposomes or microbubbles.
- 92. A composition, comprising at least a first peptide of claim 86 operatively linked to at least a first heterologous molecule.
- 93. The composition of claim 92, comprising a plurality of peptides of claim 86, wherein the peptides of the plurality are linked to each other.
- 94. The composition of claim 93, wherein peptides of the plurality are the same.
- 95. The composition of claim 92, wherein the first heterologous molecule comprises a linker moiety.
- 96. The composition of claim 95, wherein the linker moiety comprises a peptide linker.
- 97. The composition of claim 92, wherein the first heterologous molecule comprises a first heterologous peptide.
- 98. The composition of claim 97, wherein the first heterologous peptide comprises a scaffold protein.
- 99. The composition of claim 98, wherein the scaffold protein comprises a poly(amino acid).
- 100. The composition of claim 99, wherein the poly(amino acid) comprises poly(lysine).
- 101. The composition of claim 98, wherein the scaffold protein comprises a viral coat protein.
- 102. The composition of claim 101, wherein the viral coat protein comprises a Hepatitis B core protein.
- 103. The composition of claim 102, wherein the Hepatitis B core protein consists of amino acid residues 1 to 77 and amino acid residues 80 to 149 of SEQ ID NO:34, and wherein the peptide of claim 86 is operatively linked to amino acid residue 77 and to amino acid 80 of SEQ ID NO:34.
- 104. The composition of claim 101, comprising a plurality of peptides of claim 86, wherein each peptide of the plurality is operatively linked to a viral coat protein.
- 105. The composition of claim 104, which are assembled to form a virus-like particle.
- 106. The composition of claim 92, further comprising at least a second heterologous molecule.
- 107. The composition of claim 106, wherein at least the first heterologous molecule comprises a linker moiety or a scaffold protein.
- 108. The composition of claim 106, wherein the first heterologous molecule comprises a linker moiety and the second heterologous molecule comprises a scaffold protein, and wherein the linker moiety operatively links the peptide of claim NI to the scaffold protein.
- 109. The composition of claim 106, comprising, in operative linkage, a first portion of a scaffold protein, a first peptide linker, the peptide of claim N1, a second peptide linker, and a second portion of the scaffold protein.
- 110. The composition of claim 109, wherein the scaffold protein comprises a viral coat protein.
- 111. The composition of claim 92, wherein the first heterologous molecule comprises a tag.
- 112. The composition of claim 92, wherein the first heterologous molecule comprises a carrier protein.
- 113. The composition of claim 92, which is an immunogenic composition.
- 114. The composition of claim 113, further comprising an adjuvant.
- 115. A method of stimulating an immune response in a subject, comprising administering the composition of claim 92, or a polynucleotide encoding the composition, to a subject under conditions suitable for stimulating an immune response.
- 116. The method of claim 115, wherein the immune response comprises a protective immune response.
- 117. The method of claim 115, wherein the subject is a mammalian subject.
- 118. The method of claim 115, wherein the subject is a human subject.
CROSS REFERENCE TO RELATED APPLICATION(S)
[0001] This application claims the benefit of priority under 35 U.S.C. §119(e) of U.S. Serial No. 60/371,250, filed Apr. 8, 2002; U.S. Serial No. 60/371,256, filed Apr. 8, 2002; and U.S. Serial No. 60/373,668, filed Apr. 17, 2002, the entire content of each of which is incorporated herein by reference.
GRANT INFORMATION
[0002] This invention was made in part with government support under Grants Nos. 1-R43-AI52969-01 and 1-R43-C100086-01-A1 awarded by the National Institutes of Health. The government has certain rights in this invention.
Provisional Applications (3)
|
Number |
Date |
Country |
|
60371250 |
Apr 2002 |
US |
|
60371256 |
Apr 2002 |
US |
|
60373668 |
Apr 2002 |
US |