A lysine substitute for K+. A460K mutation eliminates K+ dependence in H+-pyrophosphatase of Carboxydothermus hydrogenoformans.



: Belogurov GA, Lahti R

: 2002

: Journal of Biological Chemistry

: The Journal of biological chemistry

: 277

: 51

: 4

: 0021-9258

DOI: https://doi.org/10.1074/jbc.M210341200

: http://www.ncbi.nlm.nih.gov/pubmed/12401795


The H(+) proton-translocating inorganic pyrophosphatase (H(+)-PPase) family is composed of two phylogenetically distinct types of enzymes: K(+)-dependent and K(+)-independent. However, to date, the sequence criteria governing this dichotomy have remained unknown. In this study, we describe the heterologous expression and functional characterization of H(+)-PPase from the thermophilic bacterium Carboxydothermus hydrogenoformans. Both PP(i)-hydrolyzing and PP(i)-energized H(+) translocation activities of the recombinant enzyme in Escherichia coli inner membrane vesicles are strictly K(+)-dependent. Here we deduce the K(+) requirement of all available H(+)-PPase sequences based on the K(+) dependence of C. hydrogenoformans H(+)-PPase in conjunction with phylogenetic analyses. Our data reveal that K(+)-independent H(+)-PPases possess conserved Lys and Thr that are absent in K(+)-dependent H(+)-PPases. We further demonstrate that a A460K substitution in C. hydrogenoformans H(+)-PPase is sufficient to confer K(+) independence to both PP(i) hydrolysis and PP(i)-energized H(+) translocation. In contrast, a A463T mutation does not affect the K(+) dependence of H(+)-PPase.

Last updated on 2024-26-11 at 23:22