Role of the methylcitrate cycle in propionate metabolism and detoxification in Mycobacterium smegmatis

• Published in: Microbiology (Society for General Microbiology) 2007-12, Vol.153 (12), p.3973-3982
• Main Authors: Upton, Anna M, McKinney, John D
• Format: Article
• Language: English
• Subjects: Animals; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Bacteriology; Biological and medical sciences; Carbon-Carbon Lyases - genetics; Carbon-Carbon Lyases - metabolism; Citric Acid - metabolism; Computational Biology; Culture Media; Fundamental and applied biological sciences. Psychology; Gene Deletion; Genetic Complementation Test; Isocitrate Lyase - genetics; Isocitrate Lyase - metabolism; Isoenzymes - genetics; Isoenzymes - metabolism; Metabolism. Enzymes; Mice; Microbiology; Mycobacterium smegmatis; Mycobacterium smegmatis - genetics; Mycobacterium smegmatis - growth & development; Mycobacterium smegmatis - metabolism; Mycobacterium smegmatis - pathogenicity; Mycobacterium tuberculosis; Mycobacterium tuberculosis - genetics; Propionates - metabolism; Transformation, Bacterial
• ISSN: 1350-0872; 1465-2080
• DOI: 10.1099/mic.0.2007/011726-0

Laboratory of Infection Biology, The Rockefeller University, New York, NY 10021, USA Correspondence Anna M. Upton uptona{at}rockefeller.edu Catabolism of odd-chain-length fatty acids yields acetyl-CoA and propionyl-CoA. A common pathway of propionyl-CoA metabolism in micro-organisms is the methylcitrate cycle, which includes the dedicated enzymes methylcitrate synthase (MCS), methylcitrate dehydratase (MCD) and methylisocitrate lyase (MCL). The methylcitrate cycle is essential for propionate metabolism in Mycobacterium tuberculosis . Unusually, M. tuberculosis lacks an MCL orthologue and this activity is provided instead by two isoforms of the glyoxylate cycle enzyme isocitrate lyase (ICL1 and ICL2). These bifunctional (ICL/MCL) enzymes are jointly required for propionate metabolism and for growth and survival in mice. In contrast, the non-pathogenic species Mycobacterium smegmatis encodes a canonical MCL enzyme in addition to ICL1 and ICL2. The M. smegmatis gene encoding MCL ( prpB ) is clustered with genes encoding MCS ( prpC ) and MCD ( prpD ). Here we show that deletion of the M. smegmatis prpDBC locus reduced but did not eliminate MCL activity in cell-free extracts. The residual MCL activity was abolished by deletion of icl1 and icl2 in the prpDBC background, suggesting that these genes encode bifunctional ICL/MCL enzymes. A prpB icl1 icl2 mutant was unable to grow on propionate or mixtures of propionate and glucose. We hypothesize that incomplete propionyl-CoA metabolism might cause toxic metabolites to accumulate. Consistent with this idea, deletion of prpC and prpD in the prpB icl1 icl2 background paradoxically restored growth on propionate-containing media. These observations suggest that the marked attenuation of ICL1/ICL2-deficient M. tuberculosis in mice could be due to the accumulation of toxic propionyl-CoA metabolites, rather than inability to utilize fatty acids per se. Abbreviations: DTNB, 5,5'-dithiobis-(2-nitrobenzoate); ICL, isocitrate lyase; LDH, lactate dehydrogenase; MCD, methylcitrate dehydratase; MCL, methylisocitrate lyase; MCS, methylcitrate synthase; MLS, malate synthase; PBST, PBS containing 0.05 % Tween-80 Present address : Global Health Institute, School of Life Sciences, École Polytechnique Fédérale de Lausanne (EPFL), CH-1015 Lausanne, Switzerland.