Engineering of Escherichia coli for convenient expression of ［FeFe］-hydrogenase

Abstract:

Abstract:［Objective］ A new method used to heterologously express ［FeFe］-hydrogenase in Escherichia coli was investigated in our present study． ［Methods］ By homologous recombination，three assistant genes (hydE、hydF and hydG) for hydrogenase were integrated into the chromosome of E． coli BW 25113-10，in which all hydrogenase genes were inactivated． A hydrogenase structural gene hydA from Clostridium butyricum was used to test the hydrogenase maturation ability of the recombined E． coli． BW 25113-13 ［Results］ The corrected integration of the three assistant genes was confirmed by PCR，and RT-PCR results indicated that the three accessory genes were transcripted in the recombinant． The active expression of hydA indicated that the constitutively expressed accessory proteins could assist the maturation of the ［FeFe］-hydrogenases． ［Conclusions］ A simplified ［FeFe］-hydrogenase expression recombinant E.coli BW25113-13 was constructed． It would lay foundations for the functional screening of ［FeFe］-hydrogenases and the construction of novel hydrogen producing pathways in E.coli．