Inositol monophosphate phosphatase genes of Mycobacterium tuberculosis.
BACKGROUND
Mycobacteria use inositol in phosphatidylinositol, for anchoring lipoarabinomannan (LAM), lipomannan (LM) and phosphatidylinosotol mannosides (PIMs) in the cell envelope, and for the production of mycothiol, which maintains the redox balance of the cell. Inositol is synthesized by conversion of glucose-6-phosphate to inositol-1-phosphate, followed by dephosphorylation by inositol monophosphate phosphatases (IMPases) to form myo-inositol. To gain insight into how Mycobacterium tuberculosis synthesises inositol we carried out genetic analysis of the four IMPase homologues that are present in the Mycobacterium tuberculosis genome.


RESULTS
Mutants lacking either impA (Rv1604) or suhB (Rv2701c) were isolated in the absence of exogenous inositol, and no differences in levels of PIMs, LM, LAM or mycothiol were observed. Mutagenesis of cysQ (Rv2131c) was initially unsuccessful, but was possible when a porin-like gene of Mycobacterium smegmatis was expressed, and also by gene switching in the merodiploid strain. In contrast, we could only obtain mutations in impC (Rv3137) when a second functional copy was provided in trans, even when exogenous inositol was provided. Experiments to obtain a mutant in the presence of a second copy of impC containing an active-site mutation, in the presence of porin-like gene of M. smegmatis, or in the absence of inositol 1-phosphate synthase activity, were also unsuccessful. We showed that all four genes are expressed, although at different levels, and levels of inositol phosphatase activity did not fall significantly in any of the mutants obtained.


CONCLUSIONS
We have shown that neither impA, suhB nor cysQ is solely responsible for inositol synthesis. In contrast, we show that impC is essential for mycobacterial growth under the conditions we used, and suggest it may be required in the early stages of mycothiol synthesis.
Movahedzadeh
Farahnaz
F
Department of Pathology and Infectious Diseases, Royal Veterinary College, Royal College Street, London NW1 0TU, UK. movahed@uic.edu
Wheeler
Paul R
PR
Dinadayala
Premkumar
P
Av-Gay
Yossef
Y
Parish
Tanya
T
Daffé
Mamadou
M
Stoker
Neil G
NG
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002457:Cell Extracts / Q000737:chemistry; D019295:Computational Biology; D015870:Gene Expression; D007294:Inositol / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000378:metabolism; D053858:Metabolic Networks and Pathways; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D009169:Mycobacterium tuberculosis / Q000201:enzymology* / Q000235:genetics*; D010716:Phosphatidylinositols / Q000378:metabolism; D010744:Phosphoric Monoester Hydrolases / Q000235:genetics* / Q000378:metabolism; D016415:Sequence Alignment
D001426:Bacterial Proteins; D002457:Cell Extracts; D008070:Lipopolysaccharides; D010716:Phosphatidylinositols; C050016:lipoarabinomannan; C064351:lipomannan; C034154:phosphatidylinositol mannoside; D007294:Inositol; D010744:Phosphoric Monoester Hydrolases; C021822:myo-inositol-1 (or 4)-monophosphatase