Structural Analysis And Performance Study Of Exopolysaccharides Produced By Gelidibacter Sp. PG-2

As a biological macromolecule,more and more biological application value of exopolysaccharides has been discovered,and exopolysaccharides produced by bacteria have the advantages of simple extraction and low culture cost,which have been closely paid attention by researchers.In this paper,a bacterium that can produce exopolysaccharides was found from the Bohai Sea mud,and the polysaccharides were extracted from it and the structure and properties were studied,in order to explore their application in biomedicine.In order to mine PG-2 gene-level information,this paper performed the first whole genome sequencing analysis of PG-2.The strain isolated from the Bohai Sea mud were enriched and cultured,the morphology of the strain was observed by plate scribing method and scanning electron microscopy,the strain was identified by 16S r RNA sequence analysis.the medium components of polysaccharides were optimized,the fermentation broth was extracted and purified by ethanol precipitation,trichloroacetic acid deproteinization,dialysis and column steps to obtain ZPS,and the ZPS was structurally characterized by UV,FTIR,HPLC,HPGFC,NMR,SEM,AFM and TG instruments.The effects of ZPS on cell multiplasia and migration were evaluated by MTT method and scratch experiment,and the effects of ZPS on macrophage phagocytic activity,NO release and cytokine secretion of IL-6 and TNF-αwere studied and finally the ability of ZPS as cryoprotectant and flocculant was evaluated.Whole genome sequencing of PG-2 and bioinformatics analysis.This study found that Gelidibacter sp.PG-2 secretes a novel exopolysaccharide,after optimization,the fermentation medium was determined to be LB-Z,the total sugar content of ZPS was 66.13%,and the protein content was 10.97%.ZPS is a heteropolysaccharide with a molecular weight of 492 k Da,containing mannose,glucosamine,glucuronic acid,galactoacid,aminogalactose,glucose,galactose and fucoschase,FT-IR spectroscopy showed that ZPS had a large number of hydroxyl groups and other peaks that conformed to the characteristics of polysaccharides,SEM observed that ZPS had a large pore structure,AFM analysis showed that ZPS had an irregular network structure,¹H and ¹³C NMR results showed that ZPS was an acidic polysaccharide containingαandβconfigurations,TG analysis showed that ZPS had thermal stability.Through MTT experiments,ZPS has the effect of promoting the value-added of L929 and RAW264.7,inhibiting the value-added of cancer cells A549 and MCF-7,and scratch experiments have proved that ZPS can promote the migration of normal cells L929 and inhibit the migration of cancer cells A549 and Hela.ZPS promotes phagocytic function and secretion of NO,IL-6 and TNF-αmacrophages.In addition,ZPS has proven to be cryoprotective and flocculating.The whole genome sequencing results showed that PG-2 was annotated to 3401,1797 and 1495 genes in the basic databases COG,GO and KEGG,and 180,219 and143 genes were predicted in the carbohydrate active enzyme database,virulence factor database and drug resistance gene,respectively.The new exopolysaccharide ZPS produced by Gelidibacter sp.PG-2 has the potential to be used as a tumor suppressant drug,which can be applied in medicine,cosmetics,food and other fields in the future,and the whole genome sequencing analysis has expanded the bioinformatics of this genus of bacteria,and laid a theoretical foundation for future modification of strains and customized polysaccharides.