Expression and characterization of the Escherichia coli fdo locus and a possible physiological role for aerobic formate dehydrogenase

Abstract

In the presence of nitrate, the major anaerobic respiratory pathway includes formate dehydrogenase (FDH-N) and nitrate reductase (NAR-A), which catalyze formate oxidation coupled to nitrate reduction. Two aerobically expressed isoenzymes, FDH-Z and NAR-Z, have been recently characterized. Enzymatic analysis of plasmid subclones carrying min 88 of the Escherichia coli chromosome was consistent with the location of the fdo locus encoding FDH-Z between the fdhD and fdhE genes which are necessary for the formation of both formate dehydrogenases. The fdo locus produced three proteins (107, 34, and 22 kDa) with sizes similar to those of the subunits of the purified FDH-N. In support to their structural role, these polypeptides were recognized by antibodies specific to FDH-N. Expression of a chromosomal fdo-uidA operon fusion was induced threefold by aerobic growth and about twofold by anaerobic growth in the presence of nitrate. However, it was independent of the two global regulatory proteins FNR and ArcA, which control genes for anaerobic and aerobic functions, respectively, and of the nitrate response regulator protein NARL. In contrast, a mutation affecting either the nucleoid-associated H-NS protein or the CRP protein abolished the aerobic expression. A possible role for FDH-Z during the transition from aerobic to anaerobic conditions was examined. Synthesis of FDH-Z was maximal at the end of the aerobic growth and remained stable after a shift to anaerobiosis, whereas FDH-N production developed only under anaerobiosis. Furthermore, in an fnr strain deprived of both FDH-N and NAR-A activities, aerobically expressed FDH-Z and NAR-Z enzymes were shown to reduce nitrate at the expense of formate under anaerobic conditions, suggesting that this pathway would allow the cell to respond quickly to anaerobiosis.Under anaerobic conditions and in the presence of nitrate, Escherichia coli synthesizes the major respiratory chain which, by coupling oxidation of formate to reduction of nitrate, provides the highest yield of energy available in the absence of oxygen. It is constituted by two membrane-bound protontranslocating enzymes, formate dehydrogenase N (FDH-N) and terminal nitrate reductase (NAR-A), which are linked by a quinone (46).The two corresponding fdnGHI (6) and narGHJI (8, 45) operons located at 32 and 27 min on the E. coli genetic map, respectively, have been cloned and sequenced. They encode the three ␣, ␤, and ␥ subunits of both enzymes. The ␣ subunits consist of molybdoproteins that have been proposed to contain the catalytic site. In addition, the ␣ subunit of FDH-N harbors a selenocysteine residue essential for the enzyme (5). The ␤ subunits are electron transfer units which accommodate four iron-sulfur centers, and the ␥ subunits specify the apoproteins of the b type cytochromes. Expression of the fdnGHI and narGHJI operons is induced during anaerobic growth in the presence of nitrate. It is subject to the dual control of the transcriptional activators FNR and NARL. Anaerobic induction is mediated by the g...

Analysis of the Bacterial Response to Ru(CO)3Cl(Glycinate) (CORM-3) and the Inactivated Compound Identifies the Role Played by the Ruthenium Compound and Reveals Sulfur-Containing Species as a Major Target of CORM-3 Action

“…S1, S3, and S4). The formate dehydrogenase-O genes, which rapidly increase in the expression levels in response to a switch from aerobic to anaerobic conditions, were also downregulated under these conditions (1). This indicates that the presence of CORM-3 does not simply cause the cell to shift to an anaerobic form of energy metabolism to bypass the quinol oxidases, known to be inhibited by CO (23).…”

H‐NS: a modulator of environmentally regulated gene expression

“…Two-dimensional protein gel analysis has indicated that there might be nearly as many genes whose expression is stimulated by H-NS, i.e. expression is reduced in the hns mutant, but only a few such genes have been identified and nothing is known about the mechanism by which H-NS affects their expression (Abaibou et al, 1995;Bertin et al, 1994 (vir) genes is mediated by a cascade, in which H-NS affects both production of the positive transcriptional regulator(s) of the systems and expression of the target genes of the regulator(s) (see Fig. 1 for an example).…”

The Acetate Switch

“…At neutral pH, formate is excreted. As the environmental pH decreases, depending on the availability of oxygen, formate decomposes either to carbon dioxide by aerobic formate dehydrogenase (FDO) or to both carbon dioxide and dihydrogen by formate-hydrogen lyase (FHL) (1,4,386).…”