REGULATION OF SUGAR TRANSPORT AND METABOLISM IN LACTIC ACID AND ORAL BACTERIA

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Microorganisms are causative or contributory agents in the etiology of a variety of oral diseases including dental caries, gingivitis, and periodontitis. The pathogenicity of Gram-positive streptococci and of Gram-negative anaerobes (Fusobacteria, Leptotrichia ssp.) can be attributed in part, to the generation of organic acids and toxic sulfur derivatives by these species during fermentation of dietary sugars and amino acids. The goals of this research program are two-fold. The first, is to characterize the enzymes that comprise these energy-yielding and multi-stage pathways. Second, to define the genetic and molecular mechanisms for regulation of the activities of fermentation pathways in oral bacteria. Accomplishments of the past year include : [1] , the cloning, sequence analysis, and expression in Escherichia coli of two novel enzymes (phospho-alpha- and phospho-beta-glucosidase) from Fusobacterium mortiferum; [2], the classification, sequence alignment and identification of active-site residues of enzymes constituting Group 4 of the glycosylhydrolase super-family; [3] , the characterization, mapping, cloning, and site-directed mutagenesis of the phospho-alpha- glucosidase gene (glvA) as a contribution to the Bacillus Genome Sequencing Project, and [4] , the chemical synthesis of unique chromogenic and fluorogenic phosphorylated substrates for the assay and detection of alpha- and beta- phospho-glycosylhydrolase activities. Genes encoding these unique enzymes from Fusobacteria (malH and pbgA , respectively) will be isolated and incorporated into suitable vectors. Plasmid or transposon constructs encoding these "reporter" enzymes will be used for the manipulation, and transfer of genetic information in pathogenic oral bacteria.