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Purification and functional analysis of the copper ATPase CopA of Enterococcus hirae.
Department of Clinical Pharmacology, University of Berne, 3010 Berne, Switzerland.
The Enterococcus hirae ATPase CopA is a member of the recently discovered heavy metal ATPases and shares 43% sequence identity with the human Menkes and Wilson copper ATPases. To study CopA biochemically, it was overexpressed in E. coli with an N-terminal histidine tag and purified to homogeneity by nickel affinity chromatography. The purified CopA catalyzed ATP hydrolysis with a V(max) of 0.15 micromol/min/mg and a K(m) for ATP of 0.2 mM and had an optimum pH of 6.25. The activity was 3- to 4-fold stimulated by reconstitution into proteoliposomes. The enzyme formed an acylphosphate intermediate. Its kinetics of formation and the effects of inhibitors and metal ions upon it support a function of CopA in copper transport. Purification and functional reconstitution of CopA provides the basis to study copper transport in vitro. Copyright 2001 Academic Press.
PMID: 11162579 [PubMed - indexed for MEDLINE]
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Cited by 3 PubMed Central articles
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[Biochem J. 2007]
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Induction of heavy-metal-transporting CPX-type ATPases during acid adaptation in Lactobacillus bulgaricus.
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[Appl Environ Microbiol. 2006]
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Cd2+ and the N-terminal metal-binding domain protect the putative membranous CPC motif of the Cd2+-ATPase of Listeria monocytogenes.
Bal N, Wu CC, Catty P, Guillain F, Mintz E.
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[Biochem J. 2003]