Proline Utilization by Bacillus subtilis: Uptake and Catabolism

Abstract

ABSTRACT l -Proline can be used by Bacillus subtilis as a sole source of carbon or nitrogen. We traced l -proline utilization genetically to the putBCP ( ycgMNO ) locus. The putBCP gene cluster encodes a high-affinity proline transporter (PutP) and two enzymes, the proline dehydrogenase PutB and the Δ 1 -pyrroline-5-carboxylate dehydrogenase PutC, which jointly catabolize l -proline to l -glutamate. Northern blotting, primer extension, and putB-treA reporter gene fusion analysis showed that the putBCP locus is transcribed as an l -proline-inducible operon. Its expression was mediated by a SigA-type promoter and was dependent on the proline-responsive PutR activator protein. Induction of putBCP expression was triggered by the presence of submillimolar concentrations of l -proline in the growth medium. However, the very large quantities of l -proline (up to several hundred millimolar) synthesized by B. subtilis as a stress protectant against high osmolarity did not induce putBCP transcription. Induction of putBCP transcription by external l -proline was not dependent on l -proline uptake via the substrate-inducible PutP or the osmotically inducible OpuE transporter. It was also not dependent on the chemoreceptor protein McpC required for chemotaxis toward l -proline. Our findings imply that B. subtilis can distinguish externally supplied l -proline from internal l -proline pools generated through de novo synthesis. The molecular basis of this regulatory phenomenon is not understood. However, it provides the B. subtilis cell with a means to avoid a futile cycle of de novo l -proline synthesis and consumption by not triggering the expression of the putBCP l -proline catabolic genes in response to the osmoadaptive production of the compatible solute l -proline.