Claims
- 1. A method for identifying a target for antibacterial agents, comprising determining the bacterial target of a product of a bacteriophage 44AHJD open reading frame selected from the group consisting of open reading frames 12 and 25.
- 2. The method of claim 1, wherein said determining comprises identifying at least one bacterial protein which binds to said product or a fragment thereof.
- 3. The method of claim 2, wherein said binding is determined using affinity chromatography on a solid matrix.
- 4. The method of claim 1, wherein said determining comprises identifying at least one protein:protein interaction using a genetic screen.
- 5. The method of claim 4, wherein said genetic screen is a yeast two-hybrid screen.
- 6. The method of claim 1, wherein said determining comprises a co-immunoprecipitation assay or a protein-protein crosslinking assay.
- 7. The method of claim 1, wherein said determining comprises identifying a mutated bacterial coding sequence which protects a bacterium from said bacteriophage 44AHJD open reading frame product.
- 8. The method of claim 1, wherein said determining comprises identifying a bacterial coding sequence which protects a bacterium against said product when expressed at high levels in said bacterium.
- 9. The method of claim 1, wherein said determining further comprises identifying a bacterial nucleic acid sequence encoding a polypeptide target of said product of bacteriophage 44AHJD open reading frame.
- 10. The method of claim 9, wherein said nucleic acid sequence is identified by determining at least a fragment of the amino acid sequence of a bacterial protein target, and identifying a bacterial nucleic acid sequence which encodes said fragment.
- 11. The method of claim 1, wherein said bacterial target is from an animal pathogen.
- 12. The method of claim 11, wherein said bacterial target is a gene homologous to a gene from an animal pathogen.
- 13. The method of claim 11, wherein said pathogen is a human pathogen.
- 14. The method of claim 1, wherein said bacterial target is from a plant pathogen.
- 15. The method of claim 1, wherein said bacterial target is a gene homologous to a gene from a plant pathogen.
- 16. The method of claim 1, further comprising determining the cellular or biochemical function or both of said product of bacteriophage 44AHJD open reading frame.
- 17. The method of claim 1, wherein said identifying the bacterial target comprises identifying a phage-specific site of action.
- 18. An isolated, purified, or enriched nucleic acid sequence at least 15 nucleotides in length, wherein said sequence corresponds to at least a portion of a bacteriophage 44AHJD open reading frame 12 or 25 sequence.
- 19. The nucleic acid sequence of claim 18, wherein said sequence comprises at least 50 nucleotides.
- 20. The nucleic acid sequence of claim 18, wherein said nucleic acid sequence corresponds to a fragment of said bacteriophage 44AHJD open reading frame 12 or 25 sequence.
- 21. The nucleic acid sequence of claim 20, wherein said nucleic acid sequence encodes a polypeptide which provides a bacteria-inhibiting function.
- 22. The nucleic acid sequence of claim 21, wherein said nucleic acid sequence is transcriptionally linked with regulatory sequences enabling induction of expression of said sequence.
- 23. An isolated, purified, or enriched polypeptide comprising at least a fragment of a protein encoded by Staphylococcus aureus bacteriophage 44AHJD open reading frame 12 or 25, wherein said portion is at least 5 amino acid residues in length.
- 24. The polypeptide of claim 24, wherein said polypeptide comprises a fragment at least 10 amino acid residues in length of a said polypeptide normally encoded by said bacteriophage.
- 25. A recombinant vector comprising a nucleic acid sequence at least 24 nucleotides in length corresponding to a portion of bacteriophage 44AHJD open reading frame 12 or 25.
- 26. The vector of claim 25, wherein said vector is an expression vector.
- 27. The vector of claim 26, wherein expression of said ORF is inducible.
- 28. A recombinant cell comprising a vector, wherein said vector comprises a nucleic acid sequence at least 24 nucleotides in length corresponding to at least a fragment of bacteriophage 44AHJD open reading frame 12 or 25.
- 29. The cell of claim 28, wherein said vector is an expression vector and expression of said ORF is inducible.
- 30. A method for identifying an antibacterial agent, comprising identifying an active fragment of a product of a bacteria-inhibiting ORF of a bacteriophage.
- 31. The method of claim 30, further comprising constructing a synthetic peptidomimetic molecule, wherein the structure of said molecule corresponds to the structure of said active fragment.
- 32. A method for identifying a compound active on a bacterial target protein of a bacteriophage 44AHJD open reading frame 12 or 25 product, comprising the step of contacting said bacterial target protein with a test compound; and determining whether said compound binds to or reduces the level of activity of said target protein,
wherein binding of said compound with said target protein or a reduction of the level of activity of said protein is indicative that said compound is active on said target and wherein said target is uncharacterized.
- 33. The method of claim 32, wherein said contacting is carried out in vitro.
- 34. The method of claim 32, wherein said contacting is carried out in vivo in a cell.
- 35. The method of claim 32, wherein said compound is a small molecule.
- 36. The method of claim 32, wherein said compound is a peptidomimetic compound.
- 37. The method of claim 32, wherein said compound is a fragment of a bacteriophage inhibitor protein.
- 38. The method of claim 32, further comprising determining the site of action of said compound on said target protein.
- 39. A method of screening for potential antibacterial agents, comprising the step of determining whether any of a plurality of compounds is active on a target of a bacteriophage 44AHJD open reading frame 12 or 25 product,
wherein said target is naturally produced by a pathogenic bacterium. [one step method claim]
- 40. The method of claim 39, wherein said plurality of compounds are small molecules.
- 41. A method for inhibiting a bacterium, comprising the step of;
contacting said bacterium with a compound active on a target of a bacteriophage 44AHJD open reading frame 12 or 25 product, wherein said target or target site is uncharacterized.
- 42. The method of claim 41, wherein said compound is said protein or an active fragment thereof.
- 43. The method of claim 41, wherein said compound is a structural mimetic of said protein.
- 44. The method of claim 41, wherein said compound is a small molecule.
- 45. The method of claim 41, wherein said contacting is performed in vitro.
- 46. The method of claim 41, wherein said contacting is performed in vivo in an animal.
- 47. The method of claim 41, wherein said animal is a human.
- 48. The method of claim 41, wherein said contacting is carried out in vivo in a plant.
- 49. The method of claim 41, wherein said bacterium is pathogenic.
- 50. A method for treating a bacterial infection in an animal suffering from an infection, comprising administering to said animal a therapeutically effective amount of compound active on a target of a bacteriophage 44AHJD open reading frame 12 or 25 product in a bacterium involved in said infection,
wherein said target is an uncharacterized target or the compound is active at an uncharacterized target site.
- 51. The method of claim 50, wherein said compound is a small molecule.
- 52. The method of claim 50, wherein said compound is a peptidomimetic compound.
- 53. The method of claim 50, wherein said compound is a fragment of a bacteriophage inhibitor protein.
- 54. The method of claim 50, wherein said animal is a mammal.
- 55. The method of claim 54, wherein said mammal is a human.
- 56. A method for propylactically treating an animal at risk of an infection, comprising administering to said animal a prophylactically effective amount of a compound active on a target of a bacteriophage 44AHJD open reading frame 12 or 25 product,
wherein said target is an uncharacterized target or the site of action of said compound is an uncharacterized target site.
- 57. The method of claim 56, wherein said compound is a small molecule.
- 58. The method of claim 56, wherein said compound is a peptidomimetic compound.
- 59. The method of claim 56, wherein said compound is a fragment of a bacteriophage inhibitor protein.
- 60. The method of claim 56, wherein said animal is a mammal.
- 61. The method of claim 60, wherein said mammal is a human.
- 62. An antibacterial agent active on a target of a bacteriophage 44AHJD open reading frame 12 or 25 product, wherein said target is an uncharacterized target or said agent is active at a phage-specific site on said target.
- 63. The agent of claim 62, wherein said agent is a pepetidomimetic of a bacteriophage inhibitor polypeptide.
- 64. The agent of claim 62, wherein said agent is a small molecule.
- 65. The agent of claim 62, wherein said agent is a fragment of a bacteriophage inhibitor polypeptide.
- 66. The agent of claim 62, wherein said agent is active at a phage-specific site on said target.
- 67. A method of making an antibacterial agent, comprising the steps of:
identifying a target of a bacteriophage 44AHJD open reading frame 12 or 25 product; screening a plurality of test compounds to identify a compound active on said target; and synthesizing said compound in an amount sufficient to provide a therapeutic effect when administered to an organism infected by a bacterium naturally producing said target.
- 68. The method of claim 67, wherein said compound is a small molecule.
- 69. The method of claim 67, wherein said compound is a peptidomimetic compound.
- 70. The method of claim 67, wherein said compound is a fragment or derivative of a bacteriophage 44AHJD open reading frame product.
- 71. An antibody which binds a protein encoded by an open reading frame from Staphylococcus aureus bacteriophage 44AHJD.
- 72. The antibody of claim 71, wherein said antibody binds a protein which corresponds to a protein encoded by an open reading frame from Staphylococcus aureus bacteriophage 44AHJD.
- 73. A method for detecting a phage protein comprising the steps of, contacting said phage with an antibody,
wherein said antibody binds a protein encoded by an open reading frame from Staphylococcus aureus bacteriophage 44AHJD.
- 74. A method for detecting a virus comprising the steps of, contacting said virus with an antibody,
wherein said antibody binds a protein encoded by an open reading frame from Staphylococcus aureus bacteriophage 44AHJD.
- 75. The method of claim 74, wherein said virus is pathogenic to a mammal.
- 76. The method of claim 75, wherein said mammal is a human.
- 77. A method for determining the cellular and/or biochemical function of a bacterial target of a bacteriophage 44AHJD open reading frame product comprising;
contacting said bacterial target protein with a test compound; determining whether said compound binds to or reduces the level of activity of said target protein, and identifying homologous polypeptides and/or nucleic acid molecules to said target protein having known functions, wherein binding of said compound with said target protein or a reduction of the level of activity of said protein is indicative that said compound is active on said target and wherein said target is uncharacterized.
- 78. The method of claim 77, wherein said contacting is carried out in vitro.
- 79. The method of claim 77, wherein said contacting is carried out in vivo in a cell.
- 80. The method of claim 77, wherein said compound is selected from the group consisting of a small molecule, a peptidomimetic compound, or a fragment or derivative of a bacteriophage inhibitor protein.
- 81. A method of screening for compounds that inhibit an S. aureus dnaN product, comprising
contacting said dnaN product with a bacteriophage 44AHJD ORF25 product or a fragment thereof and at least one test compound, and determining whether any of said test compounds reduces the interaction between said dnaN product and said ORF25 product, wherein a reduction in said interaction is indicative that said test compound inhibits said dnaN product.
- 82. The method of claim 81, wherein said dnaN product has the amino acid sequence of SEQ ID NO:2.
- 83. The method of claim 81, wherein said determining comprises measuring the interaction between dnaN and ORF 25 product, wherein dnaN or ORF25 product is directly labeled.
- 84. The method of claim 83, wherein said dnaN product comprises an active portion, a mimetic, a corresponding isolated, enriched, or purified protein, or a homologous product.
- 85. The method of claim 83, wherein said dnaN or ORF25 product is indirectly labeled.
- 86. The method of claim 81, wherein said detecting comprises measurement by phage display.
- 87. The method of claim 81, wherein said detecting comprises measurement by surface plasmon resonance.
- 88. The method of claim 81, wherein said detecting comprises measurement by Fluorescence Resonance Energy Transfer.
- 89. The method of claim 81, wherein said detecting comprises measurement of fluorescence polarization changes.
- 90. The method of claim 81, wherein said detecting comprises a scintillation proximity assay.
- 91. The method of claim 81, wherein said detecting comprises a biosensor assay.
- 92. The method of claim 81, wherein said test compound is a small molecule, a peptidomimetic compound, or a fragment or derivative of a bacteriophage inhibitor protein.
- 93. The method of claim 91, wherein said bacteriophage inhibitor protein is from S. aureus bacteriophage AHJD 12 or 25.
- 94. The method of claim 81, wherein said test compound is a peptide.
- 95. The method of claim 94, wherein said peptide is an artificially synthesized peptide.
- 96. The method of claim 94, wherein said peptide is a peptide prepared in expression systems.
- 97. A method for inhibiting an S. aureus dnaN product, comprising
contacting said dnaN product with a bacteriophage 44AHJD 0RF25 product or fragment thereof at a concentration sufficient to inhibit said dnaN product.
- 98. The method of claim 97, wherein said contacting is in vitro.
- 99. The method of claim 97, wherein said contacting is in a cell.
- 100. The method of claim 97, wherein said contacting is in vivo.
- 101. The method of claim 97, wherein said contacting is in a mammal.
- 102. A method for inhibiting an S. aureus dnaN product, comprising
contacting said dnaN product with a structural mimetic of a bacteriophage 44AHJD ORF25 product or biologically active fragment at a concentration sufficient to inhibit said dnaN product.
- 103. The method of claim 102, wherein said contacting is in vitro.
- 104. The method of claim 102, wherein said contacting is in a cell.
- 105. The method of claim 102, wherein said contacting is in vivo.
- 106. The method of claim 102, wherein said contacting is in a mammal.
- 107. A pharmaceutical composition comprising
a pharmaceutically effective amount of a bacteriophage 44AHJD product or fragment thereof, and a pharmaceutically acceptable carrier.
- 108. A pharmaceutical composition comprising
a pharmaceutically effective amount of a structural mimetic of a bacteriophage 44AHJD product of fragment thereof, and a pharmaceutically acceptable carrier.
- 109. The pharmaceutical composition of claim 120, wherein said structural mimetic is a peptidomimetic.
- 110. The pharmaceutical composition of claim 120, wherein said structural mimetic is a synthetic mimetic.
RELATED APPLICATIONS
[0001] This application claims the benefit of U.S. Provisional Application Pelletier, et al., No. 60/168,777 filed Dec. 1, 1999, which is hereby incorporated by reference in its entirety, including drawings.
Provisional Applications (1)
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Number |
Date |
Country |
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60168777 |
Dec 1999 |
US |