Metabolism of sugars by genetically diverse species of oral Leptotrichia

Summary Leptotrichia buccalis ATCC 14201 is a gram‐negative, anaerobic rod‐shaped bacterium resident in oral biofilm at the tooth surface. The sequenced genome of this organism reveals three contiguous genes at loci: Lebu_1525 , Lebu_ 1526 and Lebu_ 1527. The translation products of these genes exhibit significant homology with phospho‐α‐glucosidase (Pagl), a regulatory protein (GntR) and a phosphoenol pyruvate‐dependent sugar transport protein (EIICB), respectively. In non‐oral bacterial species, these genes comprise the sim operon that facilitates sucrose isomer metabolism. Growth studies showed that L. buccalis fermented a wide variety of carbohydrates, including four of the five isomers of sucrose. Growth on the isomeric disaccharides elicited expression of a 50‐kDa polypeptide comparable in size to that encoded by Lebu_1525 . The latter gene was cloned, and the expressed protein was purified to homogeneity from Escherichia coli TOP10 cells. In the presence of two cofactors, NAD + and Mn 2+ ions, the enzyme readily hydrolyzed p ‐nitrophenyl‐α‐glucopyranoside 6‐phosphate (pNPαG6P), a chromogenic analogue of the phosphorylated isomers of sucrose. By comparative sequence alignment, immunoreactivity and signature motifs, the enzyme can be assigned to the phospho‐α‐glucosidase (Pagl) clade of Family 4 of the glycosyl hydrolase super family. We suggest that the products of Lebu_1527 and Lebu_1525 , catalyze the phosphorylative translocation and hydrolysis of sucrose isomers in L. buccalis , respectively. Four genetically diverse, but 16S rDNA‐related, species of Leptotrichia have recently been described: L. goodfellowii , L. hofstadii , L. shahii and L. wadei . The phenotypic traits of these new species, with respect to carbohydrate utilization, have also been determined.