Mycothiol is essential for growth of Mycobacterium tuberculosis Erdman - PubMed (original) (raw)

Mycothiol is essential for growth of Mycobacterium tuberculosis Erdman

Dipti Sareen et al. J Bacteriol. 2003 Nov.

Abstract

Mycothiol (MSH) is the major low-molecular-mass thiol in mycobacteria and is associated with the protection of Mycobacterium tuberculosis from toxic oxidants and antibiotics. The biosynthesis of MSH is a multistep process, with the enzymatic reaction designated MshC being the ligase step in MSH production. A targeted disruption of the native mshC gene in M. tuberculosis Erdman produced no viable clones possessing either a disrupted mshC gene or reduced levels of MSH. However, when a second copy of the mshC gene was incorporated into the chromosome prior to the targeted disruption, multiple clones having the native gene disrupted and the second copy of mshC intact were obtained. These clones produced normal levels of MSH. These results demonstrate that the mshC gene and, more generally, the production of MSH are essential for the growth of M. tuberculosis Erdman under laboratory conditions.

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Figures

FIG. 1.

Key enzymes in the biosynthesis of MSH in M. tuberculosis include the glycosyltransferase (MshA, encoded by Rv0486), the GlcNAc-Ins deacetylase (MshB, encoded by Rv1170), the ATP-dependent Cys:GlcN-Ins ligase (MshC, encoded by Rv2130c), and the acetyl-CoA (CoASAc):Cys-GlcN-Ins acetyltransferase (MSH synthase; MshD, encoded by Rv0819).

FIG. 2.

Southern blot of chromosomal DNA digested with _Nco_I and probed with a 500-bp PCR fragment containing residues 195 to 708 from within the mshC gene. Erd, M. tuberculosis Erdman. Clones 59, 214, 63, 318, 317, and 331 have a single copy of mshC identical to that of the wild-type strain. Clones 151, 226, 182, and 135 have the native mshC intact and a component of the knockout substrate incorporated by nonhomologous recombination.

FIG. 3.

(A) Southern blot of chromosomal DNA digested with _Sac_I and probed with a 500-bp fragment containing residues 195 to 708 from within the mshC gene. Erd, M. tuberculosis Erdman. Clone 49 is derived from wild-type M. tuberculosis Erdman, and clones 3, 18, 157, 158, and 172 are derived from Mtb1682 (2X-mshC). (B) Diagram showing location of _Sac_I sites within the M. tuberculosis H37Rv genome near mshC and within the insert of the mshC knockout.

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Mycothiol is essential for growth of Mycobacterium tuberculosis Erdman - PubMed (original) (raw)