De novo binding domain containing polypeptides and uses thereof

Abstract

Provided herein are de novo binding domain containing polypeptides (DBDpp) that specifically bind a target of interest. Nucleic acids encoding the DBDpp, and vectors and host cells containing the nucleic acids are also provided. Libraries of DBDpp, methods of producing and screening such libraries and the DBDpp identified from such libraries and screens are also encompassed. Methods of making and using the DBDpp are additionally provided. Such uses include, without limitation, affinity purification, and diagnostic and therapeutic applications.

What is claimed is: 
     
         1 - 11 . (canceled) 
     
     
         12 . A method for transforming a reference polypeptide into a polypeptide having specific binding for a target of interest, the method comprising:
 modifying a plurality of amino acid residues from a reference polypeptide to generate a plurality of candidate binding polypeptides;
 wherein the reference polypeptide comprises a variant of a non-naturally occurring polypeptide and comprises three anti-parallel alpha helices joined by linker peptides, 
 wherein the amino acid residues to be modified are solvent accessible or solvent inaccessible, 
 wherein the modification comprises amino acid substitutions; 
   packaging the plurality of candidate binding polypeptides in a plurality of vectors to generate a candidate library; and   screening the candidate library for candidate binding polypeptides that exhibit specific binding to the target of interest.   
     
     
         13 - 17 . (canceled) 
     
     
         18 . A de novo binding domain polypeptide (DBDpp), wherein the DBDpp comprises three anti-parallel alpha helices and is a variant of a synthetic polypeptide, wherein the DBDpp immunospecifically binds to a protein that is at least 95% identical to CD137. 
     
     
         19 - 47 . (canceled) 
     
     
         48 . A method for purifying a target of interest comprising,
 contacting a sample comprising a target of interest with a composition comprising:
 a virus-like particle coupled to a solid support, wherein the virus-like particle expresses a polypeptide as a membrane protein, the polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, and SEQ ID NO:6, 
 wherein no cysteine or proline residues are substituted into any of SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, and SEQ ID NO:6, 
 wherein the polypeptide has an amino acid sequence that differs from SEQ ID NO:1, 
 wherein the polypeptide specifically binds the target of interest, 
 wherein the polypeptide's specific binding to the target of interest is greater than binding of a polypeptide according to SEQ ID NO:1 to the target of interest; and 
   the contacting performed under conditions that permit binding of the composition to the target of interest; and   removing a portion of the sample that is not bound to the composition.   
     
     
         49 - 85 . (canceled)