Application of Chimeric Glucanase of Mutanase and Dextranase for the Prevention of Dental Biofilm Formation

Objectives: Water-insoluble glucan (WIG) produced by the mutans streptococci, which is one of major cariogenic pathogens, plays an important role in the formation of dental biofilm and adhesion of the biofilm to the tooth surfaces. Glucanohydrolases, such as mutanase (α-1,3-glucanase) and dextranase (α-1,6-glucanase) are able to hydrolyze WIG. The purposes of this study were to construct bi-functional chimeric glucanase, composed of mutanase and dextranase, and to examine the effect of the chimeric glucanase on the formation and decomposition of the biofilm.
Methods: The mutanase gene from Paenibacillus humics NA1123 and the dextranase gene from Streptococcus mutans ATCC 25175 were cloned and ligated in a pE-SUMOstar Amp plasmid vector. The his-tagged fusion chimeric glucanase was expressed in Escherichia coli BL21(DE3) and partially purified. The effect of the chimeric glucanase on the formation and decomposition of the biofilm formed on glass surface by Streptococcus sobrinus 6715 GTF were examined. This biofilm was fractionated into the firm-adherent, loose-adherent and non-adherent fractions. The amounts of glucan in each fraction were determined by phenol-sulfuric acid method, and reducing sugar was determined by Somogyi-Nelson method.
Results: The chimeric glucanase reduced the formation of total amount of WIG in dose dependent manner, and statistically significant reduction of WIG of the adherent fraction was observed. Moreover, we found that the chimeric glucanase was able to decompose the formed biofilm. In consequences, the chimeric glucanase was 4.1 times more effective than the mixture of dextranase and mutanase in glucan inhibition of biofilm formation. It was suggested that the chimeric glucanase might be useful for the prevention of dental caries.
Conclusions: The chimeric glucanase composed of mutanase and dextranase effectively inhibited the biofilm formation and decomposed the biofilm.