YtsJ has the major physiological role of the four paralogous malic enzyme isoforms in Bacillus subtilis.
The Bacillus subtilis genome contains several sets of paralogs. An extreme case is the four putative malic enzyme genes maeA, malS, ytsJ, and mleA. maeA was demonstrated to encode malic enzyme activity, to be inducible by malate, but also to be dispensable for growth on malate. We report systematic experiments to test whether these four genes ensure backup or cover different functions. Analysis of single- and multiple-mutant strains demonstrated that ytsJ has a major physiological role in malate utilization for which none of the other three genes could compensate. In contrast, maeA, malS, and mleA had distinct roles in malate utilization for which they could compensate one another. The four proteins exhibited malic enzyme activity; MalS, MleA, and MaeA exhibited 4- to 90-fold higher activities with NAD+ than with NADP+. YtsJ activity, in contrast, was 70-fold higher with NADP+ than with NAD+, with Km values of 0.055 and 2.8 mM, respectively. lacZ fusions revealed strong transcription of ytsJ, twofold higher in malate than in glucose medium, but weak transcription of malS and mleA. In contrast, mleA was strongly transcribed in complex medium. Metabolic flux analysis confirmed the major role of YtsJ in malate-to-pyruvate interconversion. While overexpression of the NADP-dependent Escherichia coli malic enzyme MaeB did not suppress the growth defect of a ytsJ mutant on malate, overexpression of the transhydrogenase UdhA from E. coli partially suppressed it. These results suggest an additional physiological role of YtsJ beyond that of malate-to-pyruvate conversion.
Lerondel
Guillaume
G
Microbiologie et Génétique Moléculaire, INRA (UMR1238) CNRS (UMR2585) and INAP-G, F-78850 Thiverval-Grignon, France.
Doan
Thierry
T
Zamboni
Nicola
N
Sauer
Uwe
U
Aymerich
Stéphane
S
D001412:Bacillus subtilis / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D001426:Bacterial Proteins / Q000235:genetics / Q000502:physiology*; D003470:Culture Media; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial / Q000502:physiology; D005947:Glucose; D008291:Malate Dehydrogenase / Q000235:genetics / Q000502:physiology*; D008293:Malates / Q000378:metabolism; D005810:Multigene Family / Q000502:physiology; D009243:NAD / Q000378:metabolism; D009249:NADP / Q000378:metabolism
D001426:Bacterial Proteins; D003470:Culture Media; D008293:Malates; D009243:NAD; D009249:NADP; C030298:malic acid; D008291:Malate Dehydrogenase; D005947:Glucose