Regulation of Expression of the Fructan Hydrolase Gene of Streptococcus mutans GS-5 by Induction and Carbon Catabolite Repression

Abstract

The polymers of fructose, levan and inulin, as well as sucrose and raffinose, are substrates for the product of the fruA gene of Streptococcus mutans GS-5. The purpose of this study was to characterize the DNA immediately flanking fruA, to explore the regulation of expression of fruA by the carbohydrate source, and to begin to elucidate the molecular basis for differential expression of the gene. Located 3′ to fruA was an open reading frame (ORF) with similarity to β-fructosidases which was cotranscribed with fruA. A transcriptional initiation site, located an appropriate distance from an extended −10-like promoter, was mapped at 165 bp 5′ to the fruA structural gene. By the use of computer algorithms, two overlapping, stable stem-loop sequences with the potential to function as rho-independent terminators were found in the 5′ untranslated region. Catabolite response elements (CREs), which have been shown to govern carbon catabolite repression (CCR) by functioning as negative ciselements in gram-positive bacteria, were located close to the promoter. The levels of production of fruA mRNA and FruA were elevated in cells growing on levan, inulin, or sucrose as the sole carbohydrate source, and repression was observed when cells were grown on readily metabolizable hexoses. Deletion derivatives containing fusions of fruA promoter regions, lacking sequences 5′ or 3′ to the promoter, and a promoterless chloramphenicol acetyltransferase gene were used (i) to demonstrate the functionality of the promoter mapped by primer extension, (ii) to demonstrate that CCR of the fru operon requires the CRE that is located 3′ to the promoter region, and (iii) to provide preliminary evidence that supports the involvement of an antitermination mechanism infruA induction.

A novel signal transduction system and feedback loop regulate fructan hydrolase gene expression in Streptococcus mutans

“…Together with the real-time PCR results, these data demonstrate that the LevQRST pathway is required for the induction of EII Lev by fructose. At first, these results may seem contradictory, because, as we have described, expression of fruA in S. mutans is dramatically higher when the cells are grown in inulin or levans, than when fructose is the growth substrate (Jacques et al, 1985;Burne and Penders, 1992;Burne et al, 1999;Wen and Burne, 2002). One interpretation is that the LevQRST system binds fructose or fructose-containing polymers to mediate induction of its targets.…”

The role of fructans on dental biofilm formation by Streptococcus sobrinus, Streptococcus mutans, Streptococcus gordonii and Actinomyces viscosus

“…It has been shown that expression of the genes encoding di¡erent fructanases is tightly regulated by environmental factors [25]. Fructanase used in our study was isolated from S. mutans.…”

Characterization of two operons that encode components of fructose-specific enzyme II of the sugar:phosphotransferase system of Streptococcus mutans

“…Since the concentration of fructose provided for growth was 0.2% (11 mM) or higher, it would not be surprising to see that the fruI/fruCD double mutants still possessed signi¢cant growth capacity, even if the K m for fructose of II Man or of the EII for sorbitol was comparatively high. Note: S. mutans UA159 and its mutant strains were grown on TV medium [13] with indicated sugar as the sole carbohydrate source. PEP-dependent fructose^and glucose^PTS activities, F^PTS and G^PTS, respectively, were measured using the method of Leblanc et al [15] and expressed as nmol min 31 mg 31 cell dry wt.…”