title: Expanding the 43C9 class of catalytic antibodies using a chain-shuffling approach.
abstract: We employed a chain-shuffling technique to determine if the light chain of the catalytic antibody, 43C9, provides the best partner for the 43C9 heavy chain. Previously, we reported construction and screening of a 43C9 HC CROSS library, where the 43C9 heavy-chain gene was crossed with a library of light-chain genes in a lambda bacteriophage system. The library contained a high frequency of reconstituted antibodies recognizing the transition-state analogue. Here, we report the isolation and characterization of four of these clones. Recovered light-chain proteins share 92-96% sequence identity to the 43C9 light-chain protein. Somatic mutations of these light chains occur randomly at positions distant from the active site. Residues required for binding and catalysis were conserved. Mutations affected the topology of the binding site. Nevertheless, catalysis was not affected. Isolation of these light chains suggests the best partner for the 43C9 heavy chain is the original light chain. These clones attempt to broaden a class of 43C9-like antibodies, where the catalytic residues, His91 and Arg96, have been reproducibly selected. Similar catalytic properties between the 43C9-like antibodies suggests binding has been optimized, thus further maturation of the light chain would not lead to a better catalyst. To improve catalysis, other approaches must be considered.
authors.affiliation: Department of Chemistry, Pennsylvania State University, University Park 16802, USA.
authors.forename: G P
authors.initials: GP
authors.lastname: Miller
authors.forename: B A
authors.initials: BA
authors.lastname: Posner
authors.forename: S J
authors.initials: SJ
authors.lastname: Benkovic
mesh_terms: D000595:Amino Acid Sequence; D017151:Antibodies, Catalytic / Q000737:chemistry* / Q000235:genetics; D002384:Catalysis; D002999:Clone Cells; D006868:Hydrolysis; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D019151:Peptide Library; D013816:Thermodynamics
chemical_list: D017151:Antibodies, Catalytic; D019151:Peptide Library