Metabolic Function And Mechanism Of Chromium(?) In ChrA And ChrT Proteins

Objective: Chromium(?)metabolic-protein-coding gene ChrA and ChrT are cloned from Serratia sp.S2,and ChrA,ChrT and ChrAT prokaryotic expression vectors are constructed and transformed into E.coli BL21.By studying the characteristics of Cr(?)metabolism in engineering bacteria,to explore the roles of ChrA and ChrT genes in the function of Cr(?)metabolism and the mechanism of Cr(?)metabolism.Methods:Using Serratia sp.S2 genome as template,ChrA and ChrT genes were amplified by PCR,and expression vectors pET-28a(+)-ChrA,pET-28a(+)-ChrT and pET-28a(+)-ChrAT were constructed,and transformed into E.coli BL21 for expression.The tolerance,growth curve,reduction of Cr(?),distribution of chromium,activity of chromium reductase and intracellular oxidative stress in engineering bacteria were measured to determine the tolerance and reduction of Cr(?)in ChrA,ChrT and ChrAT engineering bacteria.To explore the tolerance and reduction of Cr(?)by ChrA,ChrT and ChrAT engineered bacteria,and analyze the mechanism of Cr(?)metabolism by engineered bacteria.Results: ChrA,ChrT and ChrAT engineering bacteria were successfully constructed by gene recombination technology.The tolerance to Cr(?): Serratia sp.S2 >ChrAT ? ChrA > ChrT> Control(P < 0.05),and the reduction ability of strains to Cr(?): Serratia sp.S2 > ChrAT?ChrT > ChrA(P < 0.05).The chromium distribution experiments confirmed that Cr(?)and Cr(?)were the main valence states.The activity of chromium reductase from electron donors involved in the reduction process of Cr(?): NADPH > NADH >non-NAD(P)H(P < 0.05).The activity of chromium reductase increasedsignificantly with NAD(P)H(P < 0.05).The Glutathione and NPSH levels of ChrA,ChrAT engineering bacteria increased significantly(P < 0.05)under the condition of Cr(?),but there was no significant difference in the index content of ChrT engineering bacteria(P > 0.05).Conclusion: ChrAT engineering bacteria possess resistance and reduction of Cr(?).It mainly reduces Cr(?)to Cr(?)by using electronic donor NAD(P)H and chrT protein as coenzyme.The reduction process promotes the production of GSH,GSSG and NPSH intracellular to maintain the reduction state.It further improves the Cr(?)tolerance and reduction ability of ChrAT engineering bacteria.