Patent ID: 6225447
Filing Date: 2001-05-01
Classification: B01J,C07K,C12N,C40B,G01N

Abstract:
A specific binding pair member which is a single chain specific binding pair member comprising a first polypeptide chain component and a second polypeptide chain component and specific for a complementary specific binding pair member of interest, produced by a method which comprises:(I) introducing into host cells;(i) first vectors comprising nucleic acid encoding a genetically diverse population of a first polypeptide chain component fused to a component of a secreted replicable genetic display package for display of said polypeptide chain component at the surface of replicable genetic display packages; and(ii) second vectors comprising nucleic acid encoding a genetically diverse population of said second polypeptide chain component;said first vectors being packaged in infectious replicable genetic display packages and their introduction into host cells being by infection into host cells harboring said second vectors; orsaid second vectors being packaged in infectious replicable genetic display packages and their introducing into host cells being by infection into host cells harboring said first vectors;(II) causing or allowing recombination between said first and second vectors within said host cells, the recombination being promoted by inclusion in said first and second vectors of sequences at which site-specific recombination occurs resulting in recombinant vectors each of which comprises nucleic acid encoding a said single chain specific binding pair member comprising a said first polypeptide chain component and a said second polypeptide chain component and an amino acid sequence encoded by a sequence provided by recombination between said sequences at which site-specific recombination occurs, and capable of being packaged into a replicable genetic display packages using said replicable genetic display package component;(III) expressing said single chain specific binding pair members within the host cells to form a library of said single chain specific binding pair members displayed by replicable genetic display packages, whereby the genetic materials of each said replicable genetic display package encodes a single chain specific binding pair member displayed at its surface,(IV) selecting by binding with said complementary specific binding pair member of interest one or more single chain specific binding pair members specific for said complementary specific binding pair member of interest, each single chain specific binding pair member thus selected being associated in its respective replicable genetic display package with nucleic acid encoding that single chain specific binding pair member,(V) obtaining nucleic acid encoding a said single chain specific binding pair member from its replicable genetic display package displaying a single chain specific binding pair member selected in step (IV);(V) producing, by expression of encoding nucleic acid in a recombinant host organism, a single chain specific binding pair member comprising a first polypeptide chain component and a second polypeptide chain component and an amino acid sequence encoded by a sequence provided by recombination between said sequences at which site-specific recombination occurs and specific for said complementary specific binding pair member of interest, which single chain specific binding pair comprises a polypeptide chain component which is as encoded by nucleic acid encoding a said polypeptide chain component of a specific binding pair member selected in step (IV) or is a derivative thereof by way of addition, deletion, substitution or insertion of one or more amino acids or by linkage of another molecule.