Journal Article

Deletion Analysis of the Subunit Genes of V-Type Na<sup>+</sup>-ATPase from <i>Enterococcus hirae</i>

Toshiaki Hosaka, Kazuma Takase, Takeshi Murata, Yoshimi Kakinuma and Ichiro Yamato

in The Journal of Biochemistry

Published on behalf of The Japanese Biochemical Society

Volume 139, issue 6, pages 1045-1052
Published in print June 2006 | ISSN: 0021-924X
Published online June 2006 | e-ISSN: 1756-2651 | DOI: http://dx.doi.org/10.1093/jb/mvj108
Deletion Analysis of the Subunit Genes of V-Type Na+-ATPase from Enterococcus hirae

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The V1Vo-ATPase from Enterococcus hirae catalyzes ATP hydrolysis coupled with sodium translocation. Mutants with deletions of each of 10 subunits (NtpA, B, C, D, E, F, G, H, I, and K) were constructed by insertion of a chloramphenicol acetyltransferase gene into the corresponding subunit gene in the genome. Measurements of cell growth rates, 22Na+ efflux activities, and ATP hydrolysis activities of the membranes of the deletion mutants indicated that V-ATPase requires nine of the subunits, the exception being the NtpH subunit. The results of Western blotting and V1-ATPase dissociation analysis suggested that the A, B, C, D, E, F, and G subunits constitute the V1 moiety, whereas the V0 moiety comprises the I and K subunits.

Keywords: ATPase activity; deletion mutant; Na+ efflux activity; subunit composition; V-type ATPase; CCCP, carbonyl cyanide m-chlorophenylhydrazone; DCCD, N,N′-dicyclohexylcarbodiimide; DTT, dithiothreitol; EDTA, ethylenediaminetetraacetic acid; KLH, keyhole limpet hemocyanin

Journal Article.  4197 words.  Illustrated.

Subjects: Biochemistry

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