On the rate of F1‐ATPase turnover during ATP hydrolysis by the single catalytic site Evidence that hydrolysis with a slow rate of product release does not occur at the alternating active site

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Abstract
Under conditions of molar excess of enzyme, isolated F1‐ATPase from beef heart mitochondria catalyses ATP hydrolysis biphasically. The rate constants for product release are ∼10−1 and 10−4−10−3 s−1, respectively. The slow phase of ATP hydrolysis is insensitive to EDTA. [γ‐32P]ATP splitting in the slow phase cannot be chased from the enzyme during several catalytic turnovers. It follows from these results that the slow single‐site hydrolysis of ATP (k cat∼10−4s−1), initially observed by Grubmeyer et al. [(1982) J. Biol. Chem. 257, 12092–12100], is not carried out by the normal catalytic site. In contrast, the phase of rapid ATP hydrolysis (k cat∼10−1s−1) is completely prevented by EDTA and is believed to be the normal function of the normal catalytic site of F1‐ATPase.