Both dextransucrase and mutansynthetase activities have been purified from the culture fluids of Streptococcus mutans GS-5 (serotype c). Although homogeneous dextransucrase preparations normally synthesize little insoluble glucan, essentially all of the glucan synthesized by this enzyme in the presence of 1.5 M (NH4)2SO4 was water insoluble. Linkage analysis of the insoluble glucans indicated that the presence of NH4' increased the portion of ca-1,3-glucose linkages relative to a-1,6-glucose units in the product. Chromatofocusing of aggregated glucosyltransferase fractions synthesizing predominantly insoluble glucan yielded primarily dextransucrase activity separable from relatively low levels of mutansynthetase activity. The latter enzyme was detected only in 18-h assays and synthesized primer-dependent insoluble glucan, which was decreased in the presence of NH4+. In the absence of primer dextran T10, the addition of dextransucrase also stimulated insoluble glucan synthesis by mutansynthetase. Dextransucrase and mutansynthetase appear to be distinct enzymes, since the latter possesses a higher molecular weight (155,000 compared to 140,000), a much lower isoelectric point, and did not cross-react with antibody directed against dextransucrase. These results are discussed relative to the mechanism of insoluble glucan synthesis by S. mutans serotype c strains.

Rough colonial morphology and bacteriocin production, two properties which may be associated with the cariogenicity of Streptococcus mutans, were transformed into several strain GS-5 mutants defective in each respective property. Transformation was determined by observing the frequency of cotransfer of these properties with different reference markers. The rough colonial transformants were identical to the parental GS-5 strain with respect to ability to synthesize water-insoluble glucans and undergo in vitro sucrose-dependent colonization of glass surfaces. Alterations in the growth medium and the concentration of the initial cell inoculum resulted in an approximate 10-fold increase in the frequency of transformation of strain GS-5 compared to previous observations.

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The antibiotic cerulenin differentially inhibited the production of glucosyltransferase activity by Streptococcus mutans GS5. Cerulenin preferentially inhibited ['4C]acetate incorporation into cellular lipids but did not affect protein synthesis or ribonucleic acid synthesis in the same manner. No significant intracellular accumulation of glucosyltransferase activity could be demonstrated in cultures treated with cerulenin. On the other hand, another inhibitor of lipid synthesis, sodium chlorophenoxyisobutyrate, did not differentially inhibit glucosyltransferase expression. In addition, the role ofa metal-requiring protease in the production of glucosyltransferase activity was suggested by the observation that the chelator quinacrine differentially inhibited the production of the enzyme.

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