Growth was performed under fermentative conditions in TGYEP, unless indicated otherwise. n. d.-not determined The results in Table 5 show that in entC or feoB mutants, expression of hyaA was reduced by approximately 50% compared with the wild type MC4100. Expression of hybO attained levels that were only approximately 10% those of hyaA (Table 5), consistent with transcriptional see more regulation data for these operons reported earlier [21]. The expression of the hybO’-'lacZ

fusion was reduced by approximately 40% in a feoB learn more mutant background and by 35% in an entC mutant compared with the level of expression measured in the wild type (Table 5). Expression of the hyc operon remained comparatively constant among the strains, but was reduced by maximally 40% in a fecA-E feoB double mutant. A slight increase in hyc expression in the feoB single mutant was observed; however, it should be noted that expression levels were variable in the mutant backgrounds. Addition

of dipyridyl to Epigenetics inhibitor the growth medium had no effect on hyc expression (data not shown). Discussion In a previous study [23] it was shown that hydrogen metabolism of E. coli was significantly affected by introduction of a fur mutation. Fur is a global regulator controlling iron homeostasis [24, 25]. Differential effects on hydrogen-oxidizing hydrogenase activity compared with hydrogen-evolving enzyme function were observed previously in the fur mutant [23]. The fur mutation, which has both negative and positive effects

on gene expression of iron metabolism including depression of iron uptake systems, caused a strong reduction in FHL activity, suggesting Fur is required for FHL synthesis. In the current study we could show in an otherwise Fur+ background that causing iron limitation by removing key iron uptake systems also resulted in differential effects on hydrogen uptake and hydrogen evolution: hydrogen-oxidizing hydrogenase function was compromised first while hydrogen-evolving hydrogenase activity was partially retained. During a search for genes affecting hydrogenase biosynthesis or activity, a mutant with a transposon insertion in feoB encoding the GTPase component of the postulated ferrous iron transport system [12] was isolated. The alteration in hydrogen metabolism oxyclozanide caused by the mutation could not be phenotypically complemented by ferrous iron but could be complemented by supplementing the growth medium with oxidized iron. This result supports the important role of the Feo system in transport of iron under reducing conditions. Although this finding was perhaps not surprising considering that the hydrogenases are synthesized under anaerobic fermentative conditions when Fe2+ ions are available and the Feo transport system is active [10–12], it was nevertheless important to demonstrate the involvement and importance of this route of iron acquisition for enzymes that have a high demand for iron atoms.