Previous Article | Next Article ![]()
Journal of Bacteriology, February 2000, p. 919-927, Vol. 182, No. 4
Department of Microbiology and Molecular
Genetics and New Jersey Medical School National Tuberculosis Center,
UMDNJ/New Jersey Medical School, Newark, New Jersey 17103
Received 22 June 1999/Accepted 19 November 1999
Genes encoding L-arginine biosynthetic and transport
proteins have been shown in a number of pathogenic organisms to be
important for metabolism within the host. In this study we describe the cloning of a gene (Rv0522) encoding an amino acid transporter from
Mycobacterium bovis BCG and the effects of its deletion on L-arginine transport and metabolism. The Rv0522 gene of BCG
was cloned from a cosmid library by using primers homologous to the rocE gene of Bacillus subtilis, a putative
arginine transporter. A deletion mutant strain was constructed by
homologous recombination with the Rv0522 gene interrupted by a
selectable marker. The mutant strain was complemented with the
wild-type gene in single copy. Transport analysis of these strains was
conducted using 14C-labeled substrates. Greatly reduced
uptake of L-arginine and
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
Amino Acid Transport and Metabolism in Mycobacteria: Cloning,
Interruption, and Characterization of an
L-Arginine/
-Aminobutyric Acid Permease in
Mycobacterium bovis BCG
-aminobutyric acid (GABA) but
not of lysine, ornithine, proline, or alanine was observed in the
mutant strain compared to the wild type, grown in Middlebrook 7H9
medium. However, when the strains were starved for 24 h or
incubated in a minimal salts medium containing 20 mM arginine (in which
even the parent strain does not grow), L-[14C]arginine uptake by the mutant but not
the wild-type strain increased strongly. Exogenous
L-arginine but not GABA, lysine, ornithine, or alanine was
shown to be toxic at concentrations of 20 mM and above to wild-type
cells growing in optimal carbon and nitrogen sources such as glycerol
and ammonium. L-Arginine supplied in the form of dipeptides
showed no toxicity at concentrations as high as 30 mM. Finally, the
permease mutant strain showed no defect in survival in unactivated
cultured murine macrophages compared with wild-type BCG.
*
Corresponding author. Mailing address: Department of
Microbiology and Molecular Genetics, UMDNJ/New Jersey Medical School, 185 South Orange Ave., Newark, NJ 07103. Phone: (973) 972-3759. Fax:
(973) 972-3644. E-mail: connell{at}umdnj.edu.
This article has been cited by other articles:
| Appl. Environ. Microbiol. | Infect. Immun. | Eukaryot. Cell |
|---|---|---|
| Mol. Cell. Biol. | J. Virol. | Microbiol. Mol. Biol. Rev. |
| ALL ASM JOURNALS |