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Membrane-bound pyrophosphatase of Thermotoga maritima requires sodium for activity
Department of Biochemistry and Food Chemistry, University of Turku, Turku, Finland; A. N. Belozersky Institute of Physico-Chemical Biology and School of Chemistry, Moscow State University, Moscow, Russia.
Department of Biochemistry and Food Chemistry, University of Turku, Turku, Finland.
A. N. Belozersky Institute of Physico-Chemical Biology and School of Chemistry, Moscow State University, Moscow, Russia.
Department of Biochemistry and Food Chemistry, University of Turku, Turku, Finland.
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2005 (English)In: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 44, no 6, 2088-2096 p.Article in journal (Refereed) Published
Abstract [en]

Membrane-bound pyrophosphatase of the hyperthermophilic bacterium Thermotoga maritima(Tm-PPase), a homologue of H(+)-translocating pyrophosphatase, was expressed in Escherichia coli and isolated as inner membrane vesicles. In contrast to all previously studied H(+)-PPases, both native and recombinant Tm-PPases exhibited an absolute requirement for Na(+) but displayed the highest activity in the presence of millimolar levels of both Na(+) and K(+). Detergent-solubilized recombinant Tm-PPase was thermostable and retained the monovalent cation requirements of the membrane-embedded enzyme. Steady-state kinetic analysis of pyrophosphate hydrolysis by the wild-type enzyme suggested that two Na(+) binding sites and one K(+) binding site are involved in enzyme activation. The affinity of the site that binds Na(+) first is increased with increasing K(+) concentration. In contrast, only one Na(+) binding site (K(+)-dependent) and one K(+) binding site were involved in activation of the Asp(703) --> Asn variant. Thus, Asp(703) may form part of the K(+)-independent Na(+) binding site. Unlike all other membrane and soluble PPases, Tm-PPase did not catalyze oxygen exchange between phosphate and water. However, solubilized Tm-PPase exhibited low but measurable PP(i)-synthesizing activity, which also required Na(+) but was inhibited by K(+). These results demonstrate that T. maritima PPase belongs to a previously unknown subfamily of Na(+)-dependent H(+)-PPase homologues and may be an analogue of Na(+),K(+)-ATPase.

Place, publisher, year, edition, pages
American Chemical Society (ACS), 2005. Vol. 44, no 6, 2088-2096 p.
National Category
Cell Biology
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URN: urn:nbn:se:liu:diva-132127DOI: 10.1021/bi048429gPubMedID: 15697234OAI: oai:DiVA.org:liu-132127DiVA: diva2:1037767
Available from: 2016-10-18 Created: 2016-10-18 Last updated: 2016-10-26Bibliographically approved

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