Characterization of Glycansucrase Activities from Leuconostoc Mesenteroides LM17 and URE 13 Strains

ABSTRACT

14 original strains from traditional Bulgarian fermented food have been studied in order to characterize new orginal glycosyltransferase activities In order to characterize the whole spectrum of functional glycosyltransferases in the studied strains, the EPS produced in liquid media where first purified and hydrolysed and the global enzymatic activities measured by DNS assay. Activities present in the culural supernatant and cell associated were determined using SD_SPAGE and in situ Periodic Acid Schiffs staining after incubation with sucrose and raffinose. Among the studied strains Lm17 and URE 13 strains shown interesting activities. Three polymers bands were detected during in situ EPS production in presence of sucrose associated respectively to ∼300 kD (only in URE 13) and 180 kDa and 120 kDa (in both Lm 17 and URE13) proteins The 180KDa band could correspond to a glucansucrase activity of these strains. Analysis of zymogram incubated in raffinose as substrate and hydrolysis of the EPS produced in liquid culture are in good correlation with the hypothesis of a fructosyltransferase activity that could be attributed to a 120 kDa extracellular and cell-associated protein‥ GTF specific motifs (YG repeats from the Glucan Binding Domain) from Lm 17 and URE 13 strains were amplified by PCR and sequenced. They share a high percentage of identity to each other, and to the YG-repeats from ATCC 8293 and NRRL B-512F. A conserved motif from the catalytic core of FTF genes from Lm 17 and URE 13 strains was also shown to be very conserved between these two strains and with sequences of levansucrase from Ln. mesenteroides NRRL B-512F.

Keywords:

• glycosyltransferase
• fructosyltransferase
• Leuconostoc mesenteroides
• GTF-gen
• FTF-gen