Robustness of the Rotary Catalysis Mechanism of F1-ATPase [Molecular Biophysics]

May 29th, 2014 by Watanabe, R., Matsukage, Y., Yukawa, A., Tabata, K. V., Noji, H.

F1-ATPase (F1) is the rotary motor protein fueled by ATP-hydrolysis. Previous studies have suggested that three charged residues are indispensable for catalysis of F1: the p-loop lysine in the phosphate-binding-loop, GXXXXGK(T/S), a glutamic acid that activates water molecules for nucleophilic attack on the γ phosphate of ATP (general base), and an arginine directly contacting the γ phosphate (arginine-finger). These residues are well conserved among p-loop NTPases. In the present study, we investigated the role of these charged residues in catalysis and torque generation by analyzing alanine-substituted mutants in the single-molecule rotation assay. Surprisingly, all mutants continuously drove rotary motion, even though the rotational velocity was at least 100,000 times slower than that of wild-type. Thus, although these charged residues contribute to highly efficient catalysis, they are not indispensable to chemo-mechanical energy coupling, and the rotary catalysis mechanism of F1 is far more robust than previously thought.