MODIFICATION OF THE MITOCHONDRIAL F1-ATPASE EPSILON-SUBUNIT, ENHANCEMENT OF THE ATPASE ACTIVITY OF THE IF1-F-1 COMPLEX AND IF1-BINDING DEPENDENCE OF THE CONFORMATION OF THE EPSILON-SUBUNIT

Treatment of bovine heart submitochondrial particles with a low concen tration of 2-hydroxy-5-nitrobenzyl bromide (HNB), a selective reagent for the Trp residue of the epsilon subunit [Baracca, Barogi, Lenaz and Solaini (1993) Int. J. Biochem. 25, 1269-1275], enhances the ATP hydr olytic activity of the particles exclusively when the natural inhibito r protein IF1 is present. Similarly, isolated F-1 [the catalytic secto r of the mitochondrial H+-ATPase complex (ATP synthase)] treated with the reagent has the ATPase activity enhanced exclusively if IF1 is bou nd to it. These experiments suggest that the modification of the epsil on subunit decreases the inhibitory activity of IF1, eliciting the sea rch for a relationship between the epsilon subunit and the inhibitory protein. Certainly, a reverse relationship exists because HNB binds co valently to the isolated F-1 exclusively when the inhibitory protein i s present. This finding is consistent with the existence of the a subu nit in different conformational states depending on whether IF1 is bou nd to F-1 or not. Support for this assertion is obtained by measuremen ts of the intrinsic phosphorescence decay rate of F-1, a probe of the Trp epsilon subunit conformation in situ [Solaini, Baracca, Parenti-Ca stelli and Strambini (1993) Eur. J. Biochem. 214, 729-734]. A signific ant difference in phosphorescence decay rate is detected when IF1 is a dded to preparations of F-1 previously devoid of the inhibitory protei n. These studies indicate that IF1 and the a subunit of the mitochondr ial F-1-ATPase complex are related, suggesting a possible role of the a subunit in the mechanism of regulation of the mitochondrial ATP synt hase.