Research On Heat Shock Response Of Rhodobacter Sphaeroides

Temperature has a profound effect on microbial evolution,which produces thermophiles,psychrophiles and psychrophile.Although the psychrophiles only normally survive at room temperature,when the temperature of the environment changes,some heat shock proteins(HSP)can be activated to ensure that the bacteria can still develop normal growth metabolism at high temperatures.These HSP are important biological resources,which have important application value in the food industry,environmental protection and medical health.Currently,the researchers have conducted extensive research on HSP,but there are still many scientific issues to explore about the initial study of the small heat shock protein(shspology).The red bacteria S1(R.sphaeroides ATCC 17023)is an important microorganism that has more applications in food and medicine,environmental protection and agriculture.In order to improve the heat resistance of the bacteria,Therefore,this thesis mainly studies the heat response proteins of the s HSP family in the R.sphaeroides S1.Firstly,R.sphaeroides S1 was heat-shocked under different temperature(30-95 ℃)that the treatment with heat shocking at 75 ℃ for 10 min was selected for the subsequent experiment.Secondly,i TRAQ proteomics analysis was used to screen the different expression proteins of S1 after shocking.COG functional enrichment analysis of these proteins showed that the expression levels of s HSP genes ibp A(RSP＿1016),htp X(RSP＿0554)and RSP＿1572,which belong to the HSP20 family,were significantly up-regulated after heat shock.Through pathway enrichment analysis,it was found that some pathways such as transcription,translation and signal transduction were related to or had a positive effect on the heat stress tolerance of S1.Among them,the fatty acid anabolic pathway was completely activated,indicating that fatty acid synthesis plays an important role in S1 resistance to heat stress.Protein interaction analysis(PPI)showed that the proteins encoded by ibp A,RSP＿1572 and htp X interacted with gro ES.Gro ES could improve the survival rate of cells under heat stress by interacting with the three proteins.In this study,we found that cold shock protein(RSP＿2346)also interacted with cytochrome c oxidoreductase Cyt o,which is one of the participants in the respiratory chain of S1.Knockout of the key subunit of Cyt o gene showed that the heat tolerance of the mutants was decreased.Combined with the above results and the existing mechanism of Gro ESL,the mechanism of s HSP in the heat shock response of R.sphaeroides S1 was preliminly constructed.Based on these findings,In order to further study the function of differential proteins in heat resistance mechanism,seven genetically engineered strains were constructed by genetic manipulation.The strains were △ibp A,ibp A/op,RSP＿1572/op,htp X/op,WT/op.△coxB-I△coxB-II and △coxB-IIcoxB-I,two key subunits of Cyt o in the respiratory chain.Finally,the biological functions of s HSP was studied.The results showed that overexpression of ibp A and RSP＿1572 could improve the survival rate of the cells under high temperature environment,while overexpression of htp X had no apparent influence on the heat resistance of the cells.The overexpression of RSP＿1572,ibp A and htp X also had a protective effect on the cells under strong acid and alkali environment,which led to a higher survival rate than the WT/op strain under highly acidic and alkaline environment.The expression of RSP＿1572 could significantly improve the survival rate of the cells under highly salty environment,and the expression of ibp A and htp X could also improve the ability of the cells to resistance to salt stress.Taken together,thw results suggested that RSP＿1572 and ibp A played important roles in the tolerance of R.sphaeroides S1 to high temperature,acid,alkali and salt stresses,while htp X played a role in the tolerance to acid,alkali and salt stresses.