Role of different Escherichia coli hydrogenases in H~+ efflux and F_1F_0-ATPase activity during glycerol fermentation at different pH values

摘要

Escherichia coli is able to ferment glycerol and produce H_2 by different Hyds (hydrogenases). Wild-type whole cells were shown to extrude H~+ through the F_1F_0-ATPase and by other means with a lower rate compared with that under glucose fermentation. At pH 7.5, H~+ efflux was stimulated in fhlA mutant (with defective transcriptional activator of Hyd-3 or Hyd-4) and was lowered in hyaB or hybC mutants (with defective Hyd-1 or Hyd-2) and hyaB hybC double mutant; DCCD (dicyclohexylcarbodi-imide)-sensitive H~+ efflux was observed. At pH 5.5, H~+ efflux in wild-type was lower compared with that at pH 7.5; it was increased in fhlA mutant and absent in hyaB hybC mutant. Membrane vesicle ATPase activity was lower in wild-type glycerol-fermented cells at pH 7.5 compared with that in glucose-fermented cells; 100 mM K~+ did not stimulate ATPase activity. The latter at pH 7.5, compared with that in wild-type, was lower in hyaB and less in hybC mutants, stimulated in the hyaB hybC mutant and suppressed in the fhlA mutant; DCCD inhibited ATPase activity. At pH 5.5, the ATPase activities of hyaB and hybC mutants had similar values and were higher compared with that in wild-type; ATPase activity was suppressed in hyaB hybC and fhlA mutants. The results indicate that during glycerol fermentation, H~+ was expelled also via F_1F_0. At pH 7.5 Hyd-1 and Hyd-2 but not FhlA or Hyd-4 might be related to F _1F_0 or have their own H~+-translocating ability. At pH 5.5, both Hyd-1 and Hyd-2 more than F_1F_0 might be involved in H~+ efflux.