Allosteric Regulation And Transcriptome-differentially Expressed Gene Analysis Of Exopolysaccharide Secreted By Pseudoalteromonas Agarivorans Under The Disturbance Of Environmental Factors

Microbial exopolysaccharide is a carbohydrate polymer produced by microorganisms and secreted to the outside of the cell.Exopolysaccharide can reduce the damage to the microorganisms caused by external environmental disturbances,and improve the chances and ability of survival.The exopolysaccharide produced by Pseudoalteromonas agarivorans Hao2018 at different temperatures and initial p H were extracted and purified.In order to analyze the influence of environmental factors on the production and structure of exopolysaccharide,we used high performance liquid chromatography(HPLC)and infrared spectroscopy(IR)to determine the composition,sugar ring form and glycosidic bond configuration of extracellular polysaccharides.The antioxidant activity of exopolysaccharide was evaluated by measuring their scavenging capacity for hydroxyl radicals,DPPH radicals and ABTS radicals under various conditions.Finally,we performed transcriptome sequencing on Pseudoalteromonas agarivorans Hao2018 at different initial p H of fermentation,and the expressions of genes related to the two-component system,bacterial chemotaxis and exopolysaccharide synthesis were analyzed.The research results of the subject are as follows:Temperature and initial p H of fermentation will affect the yield of Pseudoalteromonas agarivorans Hao2018 crude exopolysaccharide.The monosaccharide composition of exopolysaccharide is mainly glucose,mannose and rhamnose,the ratio of glucose showed a trend of increasing first and then decreasing with the increase of temperature,and decreasing with the increase of initial p H of fermentation.The trend of the ratio of mannose and rhamnose is opposite to that of glucose.IR analysis showed that the extracellular polysaccharides produced under each condition showed the characteristic absorption peaks of polysaccharides,the characteristic absorption peaks of ?-D-glucopyranose and ?-D-mannopranose.However,the exopolysaccharide produced by the initial fermentation p H9 also showed the characteristic absorption peak of ?-D-mannopranose and the characteristic absorption peak of D-furanose,indicating that the change of initial p H of fermentation will affect the exopolysaccharide ring form and glycosidic bond configuration.The exopolysaccharide produced at 25? or p H9 had higher scavenging rate to hydroxyl radicals,DPPH radicals and ABTS radicals.After comprehensive analysis,we speculate that the increase in the proportion of mannose in the monosaccharide component or the exopolysaccharide with ?-form sugar ring and furan-type glycoside bond has good antioxidant activity.Transcriptome sequencing results showed that compared with the S157 group,523 genes were up-regulated and 691 genes were down-regulated in the S159 group.The results of KEGG enrichment analysis showed that the differential genes were mainly enriched in the two-component system and the bacterial chemotaxis pathway.In the S159 group,67 genes related to the two-component system showed differential expression.Among them,the gene encoding histidine kinase Pho R(chr1?2513)and the gene Bae S(chr1?881),and the gene encoding the reaction regulator Bae R(chr1?882)were Upregulated expression,the gene encoding histidine kinase Pho Q(chr1?1805),the gene encoding response regulator protein Pho B(chr1?2514)and the gene of Rst A(chr1?933)were down-regulated.In the S159 group of genes related to bacterial chemotaxis,the expression levels of genes encoding the histidine kinase Che A(chr1?3060),genes corresponding to the response regulator Che Y(chr2?363,chr2?53),and genes corresponding to Che B(chr1?2277,chr1?3055)are higher than S157 group,Pseudoalteromonas agarivorans Hao2018 uses protein phosphorylation and dephosphorylation to regulate the flagellar motor rotation direction,so as to achieve the purpose of adapting to environmental factors.The synthesis of bacterial exopolysaccharide requires the participation of multiple types of genes including transport-related genes,nucleotide sugar synthesis-related genes,and genes encoding glycosyltransferases.The expression levels of glucose transporterrelated genes(chr1?1825,chr2?538,chr1?1082)in S159 group were higher than those in S157 group.The expression of key enzyme genes during nucleotide sugar synthesis showed that the expression levels of the genes encoding UDP-D-Glc(chr1?1334,chr1?2358,and chr1?459)in the S159 group were lower than those in the S157 group,encoding the GDP-D-Man gene the expression levels of(chr1?423 and chr1?422)in the S159 group were higher than those in the S157 group.