Two types of essential carboxyl groups in Rhodospirillum rubrum proton ATPase

Abstract

Two different types of essential carboxyl groups were detected in the extrinsic component of the proton ATPase of Rhodospirillum rubrum. Chemical modification of R. rubrum chromatophores or its solubilized ATPase by Woodward's reagent K resulted in inactivation of photophosphorylating and ATPase activities. The apparent order of reaction was nearly 1 with respect to reagent concentration and similar K 1 were obtained for the soluble and membrane-bound ATPases suggesting that inactivation was associated with modification of one essential carboxyl group located in the soluble component of the proton ATPase. Inactivation was prevented by adenine nucleotides but not by divalent cations. Dicyclohexylcarbodiimide completely inhibited the solubilized ATPase with a K 1 of 5.2 m m and a K 2 of 0.81 min −1. Mg 2+ afforded nearly complete protection with a K d of 2.8 m m. Two moles of [ 14C]dicyclohexylcarbodiimide were incorporated per mole of enzyme for complete inactivation but in the presence of 30 m m MgCl 2 only one mole was incorporated and there was no inhibition. The labeling was recovered mostly from the β subunit. The incorporation of the labeled reagent into the ATPase was not prevented by previous modification with Woodward's reagent K. It is concluded that both reagents modified two different essential carboxyl groups in the soluble ATPase from R. rubrum.