Structure Analysis And Biosynthetic Genes Of Exopolysaccharide From Lactobacillus Plantarum C88

Exopolysaccharides (EPSs) produced by lactic acid bacteria (LAB) play an important role in the improvement of physical properties of fermented dairy products. The EPS can be considered as natural biothickeners because they are produced in situ by the LAB-starters that have General Recognised As Safe status (GRAS). The physico-chemical properties of EPSs determine their viscosifying ef?ciency. A better understanding of structure-function relationships of EPS in a dairy food matrix and of EPS biosynthesis remain two major challenges for further applications of EPS. In this paper, chemical structure and biosynthesis genes of EPS from Lactobacillus plantarum C88 were studied. The investigation results were as follows:Lb. plantarum C88 is capable of producing two forms of EPS when grown in MRS broth or semi-defined medium. The capsular-polysaccharide (CPS) present surrounding the bacterial surface of Lb. plantarum C88 was observed by both optical microscopy and transmission electron microscopy. The slime-polysaccharide (SPS) present in the growth medium was produced mainly during the exponential growth phase.A neutral EPS produced by Lb. plantarum C88 was isolated from culture supernatants. The polysaccharide material was puri?ed by exchanging chromatography on DEAE-cellulose column and gel permeation chromatography on Sepharose CL-6B and its structure was determined by ion chromatography and FTIR spectroscopy. Monosaccharide analysis of the purified EPS sample showed that the EPS of Lb. plantarum C88 was composed of galactose and glucose in a molar ratio of 1:2. The molecular mass of the EPS was determined to be 1.15×106 Da. Sulfated and phosphorylated derivatives of the EPS were also prepared.To clone the CPS4A and wzb genes in Lb. plantarum C88, degenerate primers were designed from the conserved domain of the EPS biosynthetic genes of LAB. The distinctive PCR products with the expected size were amplified from Lb. plantarum C88. The upstream gene sequence(CPS4BCE, galE3 genes) of CPS4A was cloned by genomic walking, whose size was 4.9kb. Nucleotide sequence similarity analyses with GenBank sequences were performed using the BLAST software at NCBI. GenBank database searching shows that the predicted protein products of the CPS4ABCE genes, galE3 gene and wzb gene from Lb. plantarum C88 have highest identity with the EPS biosynthetic genes of Lactobacillus.