STREPTOCOCCUS-MUTANS FRUCTOSYLTRANSFERASE (FTF) AND GLUCOSYLTRANSFERASE (GTFBC) OPERON FUSION STRAINS IN CONTINUOUS CULTURE

Three glucosyltransferases (GTFs), which catalyze the formation of water-insoluble adherent glucans, and fructosyltransferase (FTF), which synthesizes fructans, are believed to contribute to the pathogenic potential of Streptococcus mutans. Study of the regulation of expression of GTF and FTF has been difficult because of the complexity and number of exoenzymes produced by this bacterium. By using continuous chemostat culture to control environmental conditions, chloramphenicol acetyltransferase (CAT) operon fusions were utilized to measure transcriptional activity of the ftf and gtfBC gene promoters. Expression of these operon fusions was differentially regulated in response to culture pH and growth rate and during transition states between growth domains. Furthermore, the addition of sucrose to steady-state cultures resulted in significant increases in CAT specific activities for both fusions. In a few cases, GTF and FTF enzyme specific activities did not parallel those of the corresponding CAT fusion activities; this lack of correspondence was likely due to posttranscriptional events controlling enzyme secretion and enzyme activity, as well as to the differential expression of dextranase(s) and fructan hydrolase by S. mutans. These results clearly demonstrate that the extracellular polymer synthesis machinery of S. mutans is regulated in a complex manner. The use of operon fusions in combination with chemostat culture is a viable approach to analyzing gene expression in S. mutans and will be helpful in defining the molecular mechanisms underlying regulation of expression of virulence attributes under conditions that may more closely mimic those in dental plaque.