Utilization of d-Ribitol by Lactobacillus casei BL23 Requires a Mannose-Type Phosphotransferase System and Three Catabolic Enzymes

• Published in: Journal of Bacteriology Vol. 195; no. 11; pp. 2652 - 2661
• Main Authors: Bourand, A, Yebra, M. J, Boël, G, Mazé, A, Deutscher, J
• Format: Journal Article
• Language: English
• Published: United States American Society for Microbiology 01.06.2013
• Subjects: Agricultural sciences; Aldehyde-Lyases - genetics; Aldehyde-Lyases - isolation & purification; Aldehyde-Lyases - metabolism; Amino Acid Sequence; Bacterial Proteins - genetics; Bacterial Proteins - isolation & purification; Bacterial Proteins - metabolism; Bacteriology; Base Sequence; Biological Transport; Carbohydrate Epimerases - genetics; Carbohydrate Epimerases - isolation & purification; Carbohydrate Epimerases - metabolism; Enzymes; Fermentation; frameshift mutation; gene deletion; Gene Expression; gene overexpression; Genes; glycolysis; Gram-positive bacteria; Lactobacillus casei; Lactobacillus casei - enzymology; Lactobacillus casei - genetics; Life Sciences; Mannose - metabolism; Metabolic Networks and Pathways; Molecular Sequence Data; mutants; Mutation; NAD - metabolism; nucleotide sequences; Operon; Pentosephosphates - metabolism; Phosphoenolpyruvate - metabolism; phosphoketolase; Recombinant Fusion Proteins; Ribitol - metabolism; Sequence Analysis, DNA; Species Specificity; Sugar Alcohol Dehydrogenases - genetics; Sugar Alcohol Dehydrogenases - isolation & purification; Sugar Alcohol Dehydrogenases - metabolism; PENTITOL METABOLISM; XYLULOSE 5-PHOSPHATE; COMPLETE GENOME SEQUENCE; D-ARABINITOL; BACILLUS-SUBTILIS; PHOSPHOKETOLASE PATHWAY; ESCHERICHIA-COLI; GENETIC-CHARACTERIZATION; NUCLEOTIDE-SEQUENCE; KLEBSIELLA-PNEUMONIAE
• ISSN: 0021-9193; 1098-5530; 1067-8832
• DOI: 10.1128/JB.02276-12

Lactobacillus casei strains 64H and BL23, but not ATCC 334, are able to ferment d-ribitol (also called d-adonitol). However, a BL23-derived ptsI mutant lacking enzyme I of the phosphoenolpyruvate:carbohydrate phosphotransferase system (PTS) was not able to utilize this pentitol, suggesting that strain BL23 transports and phosphorylates d-ribitol via a PTS. We identified an 11-kb region in the genome sequence of L. casei strain BL23 (LCABL_29160 to LCABL_29270) which is absent from strain ATCC 334 and which contains the genes for a GlpR/IolR-like repressor, the four components of a mannose-type PTS, and six metabolic enzymes potentially involved in d-ribitol metabolism. Deletion of the gene encoding the EIIB component of the presumed ribitol PTS indeed prevented d-ribitol fermentation. In addition, we overexpressed the six catabolic genes, purified the encoded enzymes, and determined the activities of four of them. They encode a d-ribitol-5-phosphate (d-ribitol-5-P) 2-dehydrogenase, a d-ribulose-5-P 3-epimerase, a d-ribose-5-P isomerase, and a d-xylulose-5-P phosphoketolase. In the first catabolic step, the protein d-ribitol-5-P 2-dehydrogenase uses NAD+ to oxidize d-ribitol-5-P formed during PTS-catalyzed transport to d-ribulose-5-P, which, in turn, is converted to d-xylulose-5-P by the enzyme d-ribulose-5-P 3-epimerase. Finally, the resulting d-xylulose-5-P is split by d-xylulose-5-P phosphoketolase in an inorganic phosphate-requiring reaction into acetylphosphate and the glycolytic intermediate d-glyceraldehyde-3-P. The three remaining enzymes, one of which was identified as d-ribose-5-P-isomerase, probably catalyze an alternative ribitol degradation pathway, which might be functional in L. casei strain 64H but not in BL23, because one of the BL23 genes carries a frameshift mutation.