Evolutionarily Divergent, Na{+}-Regulated H{+}-Transporting Membrane-Bound Pyrophosphatases

Membrane-bound PPases (mPPases) of various types consume PPi to drive active H+ or Na+ transport across membranes. H+-transporting PPases are divided into phylogenetically distinct K+-independent and K+-dependent subfamilies. Here we describe a group of 46 bacterial proteins and one archaeal protein that are only distantly related to known mPPases (23–34% sequence identity). Despite this evolutionary divergence, these proteins contain the full set of 12 polar residues that interact with PPi, the nucleophilic water, and five cofactor Mg2+ ions found in “canonical” mPPases. They also contain a specific lysine residue that confers K+ independence to canonical mPPases. Two of the proteins (from Chlorobium limicola and Cellulomonas fimi) were expressed in Escherichia coli and shown to catalyze Mg2+-dependent PPi hydrolysis coupled with electrogenic H+, but not Na+ transport, in inverted membrane vesicles. Unique features of the new H+-PPases include their inhibition by Na+ and inhibition or activation, depending on PPi concentration, by K+ ions. Kinetic analyses of PPi hydrolysis over wide ranges of cofactor (Mg2+) and substrate (Mg2PPi) concentrations indicated that the alkali cations displace Mg2+ from the enzyme, thereby arresting substrate conversion. These data define the new proteins as a novel subfamily of H+-transporting mPPases that partly r...

Source: BJ Energy - February 9, 2015 Category: Biochemistry Authors: H H Luoto, E Nordbo, A M Malinen, A A Baykov, R Lahti Tags: BJ Energy Source Type: research