Structural insight into distinct mechanisms of protease inhibition by antibodies

• DOI: 10.1073/pnas.0708251104
• Publication Date: 2007-12-06T01:59:13Z
• Authors: Yan Wu; Charles Eigenbrot; Wei-Ching Liang; Scott Stawicki; Steven Shia; Bin Fan; Rajkumar Ganesan; Michael T. Lipari; Daniel Kirchhofer
• Keywords: Immunoglobulin Fab Fragments

To better understand how the relatively flat antigen-combining sites of antibodies interact with concave shaped substrate-binding clefts proteases, we determined structures two in complex trypsin-like hepatocyte growth-factor activator (HGFA). The inhibitory antibodies, Ab58 and Ab75, were generated from a human Fab phage display library synthetic diversity three complementarity determining regions (H1, H2, H3) heavy chain, mimicking natural Ig repertoire. Biochemical studies Fab58:HGFA (3.5-A resolution) Fab75:HGFA (2.2-A complexes revealed that obstructed substrate access to active site, whereas Ab75 allosterically inhibited hydrolysis. In both cases, interacted same protruding element (99-loop), which forms part cleft. inserted its H1 H2 loops cleft occupy important interaction (S3 S2). contrast, bound at backside region corresponding thrombin exosite II, is known allosteric effector molecules. agreement structural analysis, binding assays site inhibitors enzymatic showed competitive inhibitor, partial inhibitor. These results provide insight into antibody-mediated protease inhibition. They suggest unlike canonical inhibitors, may preferentially target rim interfere catalytic machinery proteases without requiring long insertion loops.