Improvement in K-Limited Growth Rate Associated with Expression

Improvement in K-Limited Growth Rate Associated with Expression

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Unformatted text preview: J OURNAL OF BACTERIOLOGY, 0021-9193/01/$04.00 1 0 DOI: 10.1128/JB.183.11.35153520.2001 June 2001, p. 35153520 Vol. 183, No. 11 Copyright 2001, American Society for Microbiology. All Rights Reserved. NOTES Improvement in K 1-Limited Growth Rate Associated with Expression of the N-Terminal Fragment of One Subunit (KdpA) of the Multisubunit Kdp Transporter in Escherichia coli ABHIJIT A. SARDESAI 1 AND J. GOWRISHANKAR 1,2 * Centre for Cellular and Molecular Biology, Hyderabad 500 007, 1 and Centre for DNA Fingerprinting and Diagnostics, Hyderabad 500 076, 2 India Received 1 March 2001/Accepted 7 March 2001 Mutations in any one of three genes, kdpA, -B, or - C, in Escherichia coli abolish the activity of Kdp, a multisubunit K 1-ATPase that belongs to the P-type ATPase family of cation transporters. We found in this study that expression in vivo of a 135-amino-acid-long N-terminal fragment (KdpA * ), less than one-quarter the length of native KdpA, was able to mediate an improvement in K 1-limited growth rates in two different contexts, even in the absence of both KdpC and the ATPase subunit KdpB. The first context was when KdpA * was overexpressed in cells from a heterologous inducible promoter, and the second was when KdpA * was provided with a C-terminally altered extension (following a spontaneous genetic rearrangement). Our results suggest that KdpA * provides an incipient pathway for K 1 translocation which can serve to transport K 1 into the cells in response to the cytoplasmic membrane potential. The intracellular concentration of K 1 in Escherichia coli under ordinary growth conditions is around 150 mM, and there is evidence that several metabolic activities occur optimally at this concentration of K 1 . In order to cope with much lower environmental concentrations of K 1 ([K 1 ] e ), E. coli cells pos- sess several active transport systems for K 1 uptake (29). They include (i) TrkA (which has subsequently been shown to com- prise two related yet distinct uptake systems, TrkG and TrkH), (ii) TrkD (also called Kup), and (iii) Kdp, which are rendered defective in trkA, trkD, and kdp mutants, respectively. The Kdp transporter has a K m for K 1 of around 2 m M, and its synthesis is induced (at the transcriptional level) only under K 1-limiting growth conditions (reviewed in references 11 and 28). Transcriptional control of Kdp is effected by a pair of proteins, KdpD and KdpE, which constitute a dual-component regulatory system. The Kdp transporter belongs to the family of P-type ATPases (for a review, see reference 23), and com- prises four subunits (the numbers of amino acid residues are indicated in parentheses): KdpF (29), KdpA (557), KdpB (682), and KdpC (190); the proteins are encoded by the ap- propriately designated genes organized as a single operon in the order kdpFABC, with the promoter-operator region situ- ated upstream of kdpF . Mutations in any one of the genes kdpA, -B, and - C abolish Kdp transporter activity; on the other...
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This note was uploaded on 10/18/2010 for the course BIO 204 taught by Professor O'neal during the Fall '07 term at SUNY Stony Brook.

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Improvement in K-Limited Growth Rate Associated with Expression

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