H_2 Producing Activity by Escherichia Coli During Mixed Carbon Fermentation at Slightly Alkaline and Acidic pHs: Novel Functions of Hydrogenase 4 (hyf) and Hydrogenase 2 (hyb)

摘要

Molecular hydrogen (H_2) production by Escherichia coli was studied during mixed carbon (glucose and glycerol) fermentation at slightly alkaline (7.5) and acidic (5.5) pHs. Wild type cells, in the assays with glucose, produced H_2 at pH 7.5 with the same level as cells grown on glucose. Compared to wild type, H_2 production in fhlA and fhlA hyfG mutants decreased ～6.5 and ～7.9 fold, respectively. In wild type cells H_2 formation at pH 5.5 was lowered ～2 fold, compared to the cells grown on glucose. But in hyfG and hybC mutants H_2 production was decreased ～2 and ～1.6 fold, respectively. However, at pH 7.5, in the assays with glycerol, H_2 production was low, when compared to the cells grown on glycerol. In contrast to slightly alkaline pH, at pH 5.5 in the assays with glycerol H_2 production was absent. Moreover, H_2 evolution in wild type cells was inhibited by 0.3 mM N,N'-dicyclohexylcarbodiimide (DCCD), an inhibitor of the F_0F_(1-) ATPase, in a pH dependent manner. At pH 7.5 in wild type cells H_2 production was decreased ～3 fold but at pH 5.5 the inhibition was ～1.7 fold. At both pHs in fhlA mutant H_2 formation was totally inhibited by DCCD. Taken together, the results indicate that at pH 7.5 in the presence of glucose glycerol also can be fermented. They suggest that Hyd-4 mainly and Hyd-2 to some extent contribute in H_2 production by E. coli during mixed carbon fermentation at pH 5.5 whereas Hyd-1 is only responsible for H_2 oxidation.