IDENTIFICATION OF AMINO-ACID-RESIDUES IN STREPTOCOCCUS-MUTANS GLUCOSYLTRANSFERASES INFLUENCING THE STRUCTURE OF THE GLUCAN PRODUCT

The glucosyltransferases (GTFs) of mutans streptococci are important v irulence factors in the sucrose-dependent colonization of tooth surfac es by these organisms. To investigate the structure-function relations hip of the GTFs, an approach was initiated to identify amino acid resi dues of the GTFs which affect the incorporation of glucose residues in to the glucan polymer. Conserved amino acid residues were identified i n the GTF-S and GTF-I enzymes of the mutans streptococci and were sele cted for site-directed mutagenesis in the corresponding enzymes from S treptococcus mutans GS5. Conversion of six amino acid residues of the GTF-I enzyme to those present at the corresponding positions in GTF-S, either singly or in multiple combinations, resulted in enzymes synthe sizing increased levels of soluble glucans. The enzyme containing six alterations synthesized 73% water-soluble glucan in the absence of acc eptor dextran T10, while parental enzyme GTF-I synthesized no such glu can product. Conversely, when residue 589 of the GTF-S enzyme was conv erted from Thr to either Asp or Glu, the resulting enzyme synthesized primarily water-insoluble glucan in the absence of the acceptor. There fore, this approach has identified several amino acid positions which influence the nature of the glucan product synthesized by GTFs.