Methods and Compositions for Identifying D-Peptidic Compounds that Specifically Bind Target Proteins

Abstract

Methods and compositions for identifying D-peptidic compounds that specifically bind target proteins are provided. Aspects of the methods include screening libraries of 20 residue or more L-peptidic compounds for specific binding to 40 residue or more D-target proteins. Once a L-peptidic compound has been identified that specifically binds to the D-target protein, the D-enantiomer of that compound may be produced.

1 . A method comprising:
 a) contacting a 50 residue or more D-target protein with a library of 20 residue or more L-peptidic compounds that comprises compounds having a scaffold domain and a variable domain of at least 8 mutations at non-core positions of the scaffold;   b) identifying a L-peptidic compound of the library that specifically binds to the D-target protein; and   c) producing the D-peptidic compound of the identified L-peptidic compound.   
     
     
         2 . The method according to  claim 1 , wherein the 50 residue or more D-target protein is selected from the group consisting of a hormone, a growth factor, a receptor, an enzyme, a cytokine, an osteoinductive factor, a colony stimulating factor and an immunoglobulin. 
     
     
         3 . The method according to  claim 2 , wherein the 50 residue or more D-target protein is selected from the group consisting of a growth hormone, a bovine growth hormone, an insulin like growth factor, a human growth hormone, a parathyroid hormone, a proinsulin, a prorelaxin, a glycoprotein hormone, a leutinizing hormone, a hemapoietic growth factor, a fibroblast growth factor, a prolactin, a placental lactogen, a tumor necrosis factor, a mullerian inhibiting substance, an inhibin, an activin, a VEGF protein, an integrin, a RANKL protein, a NGF protein, an insulin-like growth factor-I or II, an erythropoietin, an osteoinductive factor, an interferon, a colony stimulating factor, an interleukin, an IgE protein, a bone morphogenetic protein, LIF, a SCF protein, kit-ligand, a SH2 domain containing protein, a SH3 domain containing protein, an IL-4 protein, an IL-8 protein, an apoptosis protein, a hepatocyte growth factor, a hepatocyte growth factor receptor, neutravidin, and a maltose binding protein. 
     
     
         4 . The method according to  claim 3 , wherein the D-target protein is selected from the group consisting of a VEGF protein, a RANKL protein, a NGF protein, a TNF-alpha protein, a 3BP2 protein, an ABL protein, a Src protein, an IgE protein, a BLyS protein, a PCSK9 protein, an Ang2 protein, and a  Clostridium difficile  Toxin A or B protein. 
     
     
         5 . The method according to  claim 1 , wherein the library is a phage display library. 
     
     
         6 . The method according to  claim 5 , wherein the compounds of the library comprise between 20 and 80 residues. 
     
     
         7 . The method according to  claim 6 , wherein the compounds of the library comprise between 30 and 80 residues. 
     
     
         8 . The method according to  claim 7 , wherein the compounds of the library comprise between 40 and 70 residues. 
     
     
         9 .- 10 . (canceled) 
     
     
         11 . The method according to  claim 6 , wherein the D-target protein comprises 75 or more residues. 
     
     
         12 . The method according to  claim 11 , wherein the D-target protein comprises 100 or more residues. 
     
     
         13 . The method according to  claim 12 , wherein the D-target protein comprises 125 or more residues. 
     
     
         14 .- 16 . (canceled) 
     
     
         17 . The method according to  claim 6 , wherein the compounds of the library comprise a scaffold domain and a variable domain that comprises at least 10 mutations. 
     
     
         18 . The method according to  claim 17 , wherein the variable domain comprises at least 14 mutations. 
     
     
         19 . The method according to  claim 18 , wherein the compounds of the library have at least 10 conserved residues in the scaffold domain. 
     
     
         20 . The method according to  claim 19 , wherein the library comprises 50 or more distinct L-peptidic compounds comprising a β1-β2 region and three or more different non-core mutations in a region outside of the β1-β2 region. 
     
     
         21 . The method according to  claim 1 , further comprising screening the D-peptidic compound for specific binding to the L-enantiomer of the D-target protein. 
     
     
         22 . A composition comprising:
 a 50 residue or more D-target protein; and   a 20 residue or more L-peptidic library.   
     
     
         23 . The composition according to  claim 22 , wherein:
 the 50 residue or more D-target protein is selected from the group consisting of a hormone, a growth factor, a receptor, an enzyme, a cytokine, an osteoinductive factor, a colony stimulating factor and an immunoglobulin; and   each compound of the library comprises a scaffold domain and a distinct variable domain that comprises at least 5 mutations.   
     
     
         24 . The composition according to  claim 23 , wherein the 40 residue or more D-target protein is selected from the group consisting of a VEGF protein, a RANKL protein, a NGF protein, a TNF-alpha protein, a SH2 domain-containing protein, a SH3 domain containing protein, and an IgE protein. 
     
     
         25 . The composition according to  claim 24 , wherein the library is a phage display library.