Tripartite Regulation of the Operon Involved in Glycerol Catabolism by GylR, Crp, and SigF in Mycobacterium smegmatis.

The () gene encoding glycerol-3-phosphate dehydrogenase was shown to be crucial for to utilize glycerol as the sole carbon source. The gene likely forms the operon together with and , encoding a glycerol facilitator and glycerol kinase, respectively. The () gene, whose product belongs to the IclR family of transcriptional regulators, was identified 182 bp upstream of It was demonstrated that GylR serves as a transcriptional activator and is involved in the induction of expression in the presence of glycerol. Three GylR-binding sites with the consensus sequence (GKTCGRC-N-GYCGAMC) were identified in the upstream region of by DNase I footprinting analysis. The presence of glycerol-3-phosphate was shown to decrease the binding affinity of GylR to the upstream region with changes in the quaternary structure of GylR from tetramer to dimer. Besides GylR, cAMP receptor protein (Crp) and an alternative sigma factor, SigF, are also implicated in the regulation of expression. Crp functions as a repressor, while SigF induces expression of under energy-limiting conditions. In conclusion, we suggest here that the operon is under the tripartite control of GylR, SigF, and Crp, which enables to integrate the availability of glycerol, cellular energy state, and cellular levels of cAMP to exquisitely control expression of the operon involved in glycerol metabolism. Using genetic approaches, we first revealed that glycerol is catabolized through the glycolytic pathway after conversion to dihydroxyacetone phosphate in two sequential reactions catalyzed by glycerol kinase (GlpK) and flavin adenine dinucleotide (FAD)-containing glycerol-3-phosphate dehydrogenase (GlpD) in Our study also revealed that in addition to the GylR transcriptional activator that mediates the induction of the operon by glycerol, the operon is regulated by SigF and Crp, which reflect the cellular energy state and cAMP level, respectively.