US2007032636A1PendingUtilityA1

Chimeric retroviral Gag genes and screening assays

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Assignee: SAKALIAN MICHAELPriority: Sep 28, 2000Filed: Oct 18, 2006Published: Feb 8, 2007
Est. expirySep 28, 2020(expired)· nominal 20-yr term from priority
Inventors:Michael SakalianEric Hunter
C07K 14/005C07K 2319/00C12N 2740/16222C12N 2740/16122C12N 2740/12022
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Claims

Abstract

The subject invention provides novel and advantageous methods for identifying amino acid sequences in random peptide libraries that can bind to Gag polypeptides. The subject invention also establishes a novel in vitro system that can be used to test competitive inhibitors of retroviral capsid assembly. Also provided are peptides, and compositions containing these peptides, which are inhibitors of the retrovirus Gag protein(s) function. Chimeric Gag polypeptides are also provided.

Claims

exact text as granted — not AI-modified
1 . An isolated chimeric Gag polypeptide comprising at least a portion of the Mason-Pfizer Monkey Virus (M-PMV) p12 domain and at least a portion of another retroviral Gag polypeptide, said portion of said M-PMV p12 domain inducing the spontaneous assembly of said chimeric Gag polypeptide into viral capsids.  
     
     
         2 . The isolated chimeric Gag polypeptide according to  claim 1 , wherein said retroviral Gag polypeptide is obtained from human immunodeficiency virus (HIV).  
     
     
         3 . The isolated chimeric Gag polypeptide according to  claim 2 , wherein said human immunodeficiency virus is HIV-1.  
     
     
         4 . The isolated chimeric Gag polypeptide according to  claim 2 , wherein said human immunodeficiency virus is HIV-2.  
     
     
         5 . The isolated chimeric Gag polypeptide according to  claim 1 , further comprising at least one tag.  
     
     
         6 . The isolated chimeric Gag polypeptide according to  claim 5 , wherein said tag is a polyhistidine tag, a biotin tag, or a combination of polyhistidine and biotin tags.  
     
     
         7 . The isolated chimeric Gag polypeptide according to  claim 1 , further comprising a biotinylation recognition sequence (Avitag).  
     
     
         8 . A composition comprising a carrier and an isolated chimeric Gag polypeptide comprising at least a portion of the Mason-Pfizer Monkey Virus (M-PMV) p12 domain and at least a portion of another retroviral Gag polypeptide, said portion of said M-PMV p12 domain inducing the spontaneous assembly of said chimeric Gag polypeptide into viral capsids.  
     
     
         9 . An isolated polynucleotide comprising a polynucleotide sequence encoding a chimeric Gag polypeptide comprising at least a portion of the Mason-Pfizer Monkey Virus (M-PMV) p12 domain and at least a portion of another retroviral Gag polypeptide, said portion of said M-PMV p12 domain inducing the spontaneous assembly of said chimeric Gag polypeptide into viral capsids.  
     
     
         10 . The isolated polynucleotide according to  claim 9 , wherein said polynucleotide further comprises control elements.  
     
     
         11 . The isolated polynucleotide according to  claim 9 , wherein said polynucleotide further comprises a vector.  
     
     
         12 . The isolated polynucleotide according to  claim 9 , further comprising a polynucleotide sequence encoding at least one tag.  
     
     
         13 . The isolated polynucleotide according to  claim 12 , wherein said tag encoding polynucleotide sequence encodes a polyhistidine tag, a biotinylation recognition sequence, or combinations thereof.  
     
     
         14 . A host cell transformed with an isolated polynucleotide comprising a polynucleotide sequence encoding a chimeric Gag polypeptide comprising at least a portion of the Mason-Pfizer Monkey Virus (M-PMV) p12 domain and at least a portion of another retroviral Gag polypeptide, said portion of said M-PMV p12 domain inducing the spontaneous assembly of said chimeric Gag polypeptide into viral capsids.  
     
     
         15 . A method of screening for retroviral capsid assembly inhibitors comprising: 
 a. contacting one or more chimeric Gag polypeptides with candidate inhibitory compounds, said chimeric Gag polypeptide comprising at least a portion of the Mason-Pfizer Monkey Virus (M-PMV) p12 domain and at least a portion of another retroviral Gag polypeptide, said portion of said M-PMV p12 domain inducing the spontaneous assembly of said chimeric Gag polypeptide into viral capsids; and    b. screening for the presence or absence of retroviral capsid assembly.    
     
     
         16 . The method according to  claim 15 , wherein said retroviral Gag polypeptide is obtained from human immunodeficiency virus (HIV).  
     
     
         17 . The method according to  claim 16 , wherein said human immunodeficiency virus is HIV-1.  
     
     
         18 . The method according to  claim 16 , wherein said human immunodeficiency virus is HIV-2.  
     
     
         19 . The method according to  claim 15 , wherein said chimeric Gag polypeptide comprises Ch1, Ch2, Ch3, Ch3a, Ch4, or Ch7.  
     
     
         20 . The method according to  claim 15 , wherein said candidate inhibitory compounds are selected from the group consisting of peptides, peptide libraries, combinatorial peptide libraries, phage display peptide libraries, random peptide expression libraries, and synthetic peptide combinatorial libraries.  
     
     
         21 . The method according to  claim 15 , wherein said screening is conducted in an in vitro transcription and translation system.  
     
     
         22 . The method according to  claim 15 , wherein said screening is conducted in a yeast two-hybrid system.  
     
     
         23 . The method according to  claim 15 , wherein said screening is conducted in a reticulocyte lysate system.

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