Induction and Transcription Studies of the Dextransucrase Gene in Leuconostoc mesenteroides NRRL B-512F

Abstract

Dextransucrase production by Leuconostoc mesenteroidesNRRL B-512F in media containing carbon sources other than sucrose is reported for the first time. Dextransucrases were analyzed by gel electrophoresis and by an in situ activity assay. Their polymers and acceptor reaction products were also compared by 13C nuclear magnetic resonance and high-performance liquid chromatography techniques, respectively. From these analyses, it was found that, independently of the carbon source, L. mesenteroides NRRL B-512F produced dextransucrases of the same size and product specificity. The 5′ ends of dextransucrase mRNAs isolated from cells grown under different culture conditions were identical. Based on this evidence, we conclude that dextransucrases obtained from cells grown on the various carbon sources result from the transcription of the same gene. The control of expression occurs at this level. The low dextransucrase yields from cultures in d-glucose ord-fructose and the enhancement of dextransucrase gene transcription in the presence of sucrose suggest that an activating phenomenon may be involved in the expression mechanism. Dextransucrase mRNA has a size of approximately 4.8 kb, indicating that the gene is located in a monocistronic operon. The transcription start point was localized 34 bp upstream from the ATG start codon. The −10 and −35 sequences found, TATAAT and TTTACA, were highly homologous to the only glycosyltransferase promoter sequence reported for lactic acid bacteria.

Dextran production by Lactobacillus sakei MN1 coincides with reduced autoagglutination, biofilm formation and epithelial cell adhesion

“…Fructose accumulation has been previously described for dextran producing Lc. mesenteroides strains (Dols, Chraibi, Remaud-Simeon, Lindley & Monsan, 1997;Quirasco, López-Munguía, Remaud-Simeon, Monsan & Farrés, 1999;Santos, Teixeira & Rodrigues, 2000).…”

Proteolytic processing of dextransucrase of Leuconostoc mesenteroides

“…The di¤culties in identifying a protease in DS preparations allowed the hypothesis of the existence of 57^62 kDa subunits of DS [5^7]. It has recently been reported that the L. mesenteroides B-512F DS gene is located in a monocistronic operon [19]. The L. mesenteroides NRRL B-512F DS gene sequence encodes 1527 amino acids including the signal peptide [20], resulting in a predicted 170 kDa protein, that upon elimination of the signal peptide would be reduced to 166 kDa.…”

Screening and characterization of extracellular polysaccharides produced by Leuconostoc kimchii isolated from traditional fermented pulque beverage

“…For this purpose, the soluble GTF fraction was precipitated from the supernatant cultures by the addition of one volume of 50% (w/v) polyethylene glycol 5000 and centrifugation (7,000 × g , 5 min at 4°C). The pellet of cells containing the cell-associated GTF fraction, was washed three times and resuspended in a minimal volume of 0.1 mM, pH 5.4 acetate buffer (Quirasco et al 1999 ). Polymer was produced in both fractions in the same buffer conditions containing 10% sucrose in final volume of 600 μL and incubated at 30°C and 100 rpm for 14 h (New Brunswick Scientific G24 shaker).…”