Protein scaffolds for antibody mimics and other binding proteins

Abstract

Disclosed herein are proteins that include a fibronectin type III domain having at least one randomized loop. Also disclosed herein are nucleic acids encoding such proteins and the use of such proteins in diagnostic methods and in methods for evolving novel compound-binding species and their ligands.

1 - 67 . (canceled)  
     
     
         68 . A protein comprising: 
 (a) a first fibronectin type III (Fn3) domain wherein said first Fn3 domain has an amino acid sequence that has been altered relative to the amino acid sequence of a first naturally occurring Fn3 domain such that the first Fn3 domain binds to a first compound that is not bound by the first naturally occurring Fn3 domain; and    (b) a second Fn3 domain, wherein said second Fn3 domain has an amino acid sequence that has been altered relative to the amino acid sequence of a second naturally occurring Fn3 domain such that the second Fn3 domain binds to a compound that is not bound by the second naturally occurring Fn3 domain.    
     
     
         69 . The protein of  claim 68 , wherein said protein further comprises one or more additional Fn3 domains.  
     
     
         70 . The protein of  claim 69 , wherein each of said additional Fn3 domains has an amino acid sequence that has been altered relative to the amino acid sequence of a naturally occurring Fn3 domain such that each said additional Fn3 domain binds to a compound that is not bound by the corresponding naturally occurring Fn3 domain.  
     
     
         71 . The protein of  claim 68 , wherein said first Fn3 domain has an amino acid sequence that is altered relative to the amino acid sequence of a first human Fn3 domain, such that the first Fn3 domain binds to a first compound that is not bound by the first human Fn3 domain.  
     
     
         72 . The protein of  claim 71 , wherein said first Fn3 domain has an amino acid sequence that is at least 50% identical to the amino acid sequence of said first human Fn3 domain.  
     
     
         73 . The protein of  claim 71 , wherein said first human Fn3 domain is an Fn3 domain of fibronectin.  
     
     
         74 . The protein of  claim 73 , wherein said first human Fn3 domain is the tenth Fn3 module of human fibronectin ( 10 Fn3).  
     
     
         75 . The protein of  claim 68 , wherein the first and second Fn3 domains are altered in one or more loops relative to the first or second naturally occurring Fn3 domains, respectively.  
     
     
         76 . The protein of  claim 75 , wherein the first and second Fn3 domains are altered in two or more loops relative to the first or second naturally occurring Fn3 domains, respectively.  
     
     
         77 . The protein of  claim 74 , wherein the amino acid sequence of the first Fn3 domain is altered relative to the amino acid sequence of human 10Fn3 in one or more loops defined by amino acids 21-31, 51-6 and 76-88 in human Fn3.  
     
     
         78 . The protein of  claim 77 , wherein the amino acid sequence of the first Fn3 domain is altered relative to the amino acid sequence of human  10 Fn3 in one or more beta strands.  
     
     
         79 . The protein of  claim 73 , wherein said second Fn3 domain has an amino acid sequence that is altered relative to the amino acid sequence of a second human Fn3 domain, such that the second human Fn3 domain binds to a second compound that is not bound by the second human Fn3 domain.  
     
     
         80 . The protein of  claim 79 , wherein said second Fn3 domain has an amino acid sequence that is at least 50% identical to the amino acid sequence of said second human Fn3 domain.  
     
     
         81 . The protein of  claim 80 , wherein said second human Fn3 domain is an Fn3 domain of fibronectin.  
     
     
         82 . The protein of  claim 81 , wherein said second human Fn3 domain is the tenth Fn3 module of human fibronectin ( 10 Fn3).  
     
     
         83 . The protein of  claim 68 , wherein said first and second naturally occurring Fn3 domains are the same.  
     
     
         84 . The protein of  claim 68 , wherein said first and second compounds are the same.  
     
     
         85 . The protein of  claim 68 , wherein said first and second compounds are different.  
     
     
         86 . The protein of  claim 68 , wherein said first compound is a first protein.  
     
     
         87 . The protein of  claim 86 , wherein said second compound is a second protein.  
     
     
         88 . The protein of  claim 69 , wherein said protein comprises: 
 (a) a first Fn3 domain wherein said first Fn3 domain has an amino acid sequence that has been altered relative to the amino acid sequence of the eighth Fn3 module of human fibronectin ( 8 Fn3) such that the first Fn3 domain binds to a first compound that is not bound by human  8 Fn3;    (b) a second Fn3 domain wherein said first Fn3 domain has an amino acid sequence that has been altered relative to the amino acid sequence of the ninth Fn3 module of human fibronectin (9Fn3) such that the second Fn3 domain binds to a second compound that is not bound by human  9 Fn3; and    (c) a third Fn3 domain wherein said third Fn3 domain has an amino acid sequence that has been altered relative to the amino acid sequence of the tenth Fn3 module of human fibronectin ( 10 Fn3) such that the third Fn3 domain binds to a first compound that is not bound by human  10 Fn3.