Rotation of nucleotide sites is not required for the enzymatic activity of chloroplast coupling factor 1.
New heterobifunctional photoaffinity cross-linking reagents, 6-maleimido-N-(4-benzoylphenyl)hexanamide, 12-maleimido-N-(4-benzoylphenyl)dodecanamide, and 12-[14C]maleimido-N-(4-benzoylphenyl)dodecanamide, were synthesized to investigate the mechanism of ATP hydrolysis by chloroplast coupling factor 1. These reagents react with sulfhydryl groups on the gamma-polypeptide. Subsequent photolysis cross-links the gamma-polypeptide covalently to alpha- and beta-polypeptides. The cross-linkers prevent major movements of the gamma-polypeptide with respect to the alpha- and beta-polypeptides but are sufficiently long to permit some flexibility in the enzyme structure. When approximately 50% of the gamma-polypeptide was cross-linked to alpha- and beta-polypeptides, a 7% loss in ATPase activity was observed for the longer cross-linker and a 12% loss for the shorter. These results indicate that large movements of alpha- and beta-polypeptides with respect to the gamma-polypeptide are not essential for catalysis. In particular, rotation of the polypeptide chains to create structurally equivalent sites during catalysis is not a required feature of the enzyme mechanism.
D001665:Binding Sites; D003432:Cross-Linking Reagents / Q000138:chemical synthesis* / Q000494:pharmacology; D007202:Indicators and Reagents; D010944:Plants / Q000201:enzymology*; D006180:Proton-Translocating ATPases / Q000378:metabolism*; D013329:Structure-Activity Relationship
D003432:Cross-Linking Reagents; D007202:Indicators and Reagents; D006180:Proton-Translocating ATPases