Growth of Bacillus subtilis on citrate and isocitrate is supported by the Mg-citrate transporter CitM

Bacillus subtilis 168 was assayed for growth on TCA cycle intermediates and related compounds as sole carbon source. Growth was supported by citrate, D-isocitrate, succinate, fumarate and L-malate, whereas no growth was observed in the presence of cis-aconitate, αketoglutarate, D-malate, oxaloacetate and tricarballylate. Growth on the tricarboxylates citrate and D-isocitrate required the presence of functional CitM, a Mg-citrate transporter, while growth on succinate, fumarate and L-malate appeared to be CitM-independent. Interestingly, the naturally occurring D-isocitrate was favored over L-isocitrate. Like citrate, D-isocitrate was shown to be an inducer of citM expression. The addition of 1 mM Mg to the growth medium improved growth on both citrate and D-isocitrate suggesting that Disocitrate was taken up by CitM in complex with divalent metal ions. Subsequently, the ability of CitM to transport D-isocitrate was demonstrated by competition experiments and by heterologous exchange in RSO membrane vesicles prepared of E. coli cells expressing the citM gene. None of the other TCA cycle intermediates and related compounds tested was recognized by CitM. Uptake experiments using radioactive Ni provided direct evidence that D-isocitrate is transported in complex with divalent metal ions.