Mechanism of water‐insoluble glucan synthesis in Streptococcus sobrinus

Abstract

Synthesis of water-insoluble glucan (IG) by 1,3-alpha-D-glucan synthase from Streptococcus sobrinus was examined using methylation analysis. The purified enzyme was incubated with sucrose and dextran T2000 (DT2000) for a given time and only IG was harvested by centrifugation. The remaining supernatant was incubated again, and IG was obtained. By repeating the above method using the residual supernatant, we obtained 5 varieties of IG precipitated in different periods. These IGs were methylated and examined using gas chromatograph mass spectrometry. The DT2000 water-insolubilized in the early reaction stage tended to have a highly ramified structure, with 1,3-alpha-D-glucan on a 1,6-alpha-D-glucan chain as the main chain. On the contrary, the DT2000 water-insolubilized in the late stage tended to have sparse side chains of 1,3-alpha-D-glucan which elongated with incubation. Specifically, the greater the number of side chains, the sooner the DT2000 was insolubilized. These results suggest that water-insolubilization of the water-soluble glucan not only depends on the increase of the ratio of 1,3-alpha-glucoside linkages to 1,6-alpha-glucoside linkages but also on the degree of branching of the 1,3,6-alpha-branched glucoside linkages.

Effect of antiplaque compounds and mouthrinses on the activity of glucosyltransferases from Streptococcus sobrinus and insoluble glucan production

“…In the growth conditions used, S. sobrinus ATCC 33478 produced at least three glucansucrases that (i) were co‐purified by ammonium sulfate precipitation and (ii) had sufficient primer‐independent activity to produce a detectable glucan on SDS–PAGE (dextran T10 did not diffuse in the gel). The recovery level obtained was very high compared to those described by others (19, 49, 54). The molecular weights of the GTFs were in the range of 160–175 kDa generally described for this family of enzymes (8).…”

“…and Takehara et al. (26, 49) showed that the hydrolysis of low concentration sucrose and in the absence of dextran T10 was characteristic of S. sobrinus GTF, and especially GTF‐I. Sucrose hydrolysis was promoted at low sucrose concentrations.…”

Effect of inactivation of gtf genes on adherence of Streptococcus downei

“…The apparent increase in soluble glucan synthesis can be explained by considering the proposed mechanism of insoluble glucan synthesis. It is thought that GTF‐I attaches to a primer or acceptor molecule and synthesizes a 1,3‐α‐linked chain, which leads to the insolubilization of the primer ( 19, 35, 37). The minimum size of the primer is 10 glucose residues ( 23), and the glucan of GTF‐S, which averages 25 residues, seems likely to be a good source of primer.…”

Four glucosyltransferases, GtfJ, GtfK, GtfL and GtfM, from Streptococcus salivarius ATCC 25975

“…The GTF-Is synthesize chains of ɑ-(l → 3)-linked glucosyl residues onto the ɑ-(l → 6)-linked backbone of dextrans to form insoluble mixed-linkage glucans. The insolubility of these glucans is due to the ɑ-(1 → 3)-linked glucosyl residues as well as the degree of branching of the final glucan product (Walker, 1978; Walker & Jacques, 1987; Takehara et al, 1992).…”