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Structural basis for amide hydrolysis catalyzed by the 43C9 antibody
M M Thayer1, E H Olender, A S Arvai
1Department of Molecular Biology, The Scripps Research Institute, La Jolla, CA, 92037, USA.
Journal of Molecular Biology
|August 10, 1999
Summary
The antibody 43C9 uniquely catalyzes amide hydrolysis through nucleophilic attack by His L91. Crystallographic structures reveal its deep binding site and a network stabilizing the reaction transition state.
Area of Science:
- Biochemistry
- Structural Biology
- Immunology
Background:
- Catalytic antibodies offer unique reaction pathways.
- Antibody 43C9 is known for selective amide hydrolysis, a challenging reaction.
Purpose of the Study:
- To elucidate the structural and mechanistic basis of antibody 43C9's catalytic activity.
- To provide crystallographic evidence supporting computational and biochemical findings.
Main Methods:
- X-ray crystallography of antibody 43C9 single-chain variable fragment (scFv) with and without product.
- Analysis of structural features, hydrogen-bonding networks, and residue positioning.
Main Results:
- Revealed an extended beta-sheet in the heavy chain's third complementarity-determining region (CDR-H3).
- Identified a deep antigen-binding site with a hydrophobic pocket.
- Confirmed the roles of His L91 (nucleophile) and Arg L96 (stabilization) and a water-mediated hydrogen-bonding network.
Conclusions:
- The unique structure of 43C9, including the CDR-H3 conformation and deep binding site, facilitates amide hydrolysis.
- Nucleophilic attack by His L91, coupled with transition-state stabilization, drives the antibody's high catalytic efficiency.
- Water molecules in the active site may mimic the antigen transition state, enhancing catalysis.