Format

Send to

Choose Destination
Biochemistry. 2007 Jul 31;46(30):8872-8. Epub 2007 Jul 3.

Na+-pyrophosphatase: a novel primary sodium pump.

Author information

1
Department of Biochemistry and Food Chemistry, University of Turku, FIN-20014 Turku, Finland.

Abstract

Membrane-bound pyrophosphatase (PPase) is commonly believed to couple pyrophosphate (PPi) hydrolysis to H+ transport across the membrane. Here, we demonstrate that two newly isolated bacterial membrane PPases from the mesophile Methanosarcina mazei (Mm-PPase) and the moderate thermophile Moorella thermoacetica and a previously described PPase from the hyperthermophilic bacterium Thermotoga maritima catalyze Na+ rather than H+ transport into Escherichia coli inner membrane vesicles (IMV). When assayed in uncoupled IMV, the three PPases exhibit an absolute requirement for Na+ but display the highest hydrolyzing activity in the presence of both Na+ and K+. Steady-state kinetic analysis of PPi hydrolysis by Mm-PPase revealed two Na+ binding sites. One of these sites can also bind K+, resulting in a 10-fold increase in the affinity of the other site for Na+ and a 2-fold increase in maximal velocity. PPi-driven 22Na+ transport into IMV containing Mm-PPase was unaffected by the protonophore carbonyl cyanide m-chlorophenylhydrazone, inhibited by the Na+ ionophore monensin, and activated by the K+ ionophore valinomycin. The Na+ transport was accompanied by the generation of a positive inside membrane potential as reported by Oxonol VI. These findings define Na+-dependent PPases as electrogenic Na+ pumps. Phylogenetic analysis suggests that ancient gene duplication preceded the split of Na+- and H+-PPases.

PMID:
17605473
DOI:
10.1021/bi700564b
[Indexed for MEDLINE]

Supplemental Content

Full text links

Icon for American Chemical Society
Loading ...
Support Center