In Escherichia coli the expression of the nuo genes encoding the proton pumping NADH dehydrogenase I is stimulated by the presence of fumarate during anaerobic respiration. The regulatory sites required for the induction by fumarate, nitrate and 0, are located at positions around -309, -277, and downstream of -231 bp, respectively, relative to the transcriptional-start site. The fumarate regulator has to be different from the 0, and nitrate regulators ArcA and NarL. For growth by fumarate respiration, the presence of NADH dehydrogenase I was essential, in contrast to aerobic or nitrate respiration which used preferentially NADH dehydrogenase 11. The electron transport from NADH to fumarate strongly decreased in a mutant lacking NADH dehydrogenase I. The mutant used acetyl-CoA instead of fumarate to an increased extent as an electron acceptor for NADH, and excreted ethanol. Therefore, NADH dehydrogenase I is essential for NADH + fumarate respiration, and is able to use menaquinone as an electron acceptor. NADH + dimethylsulfoxide respiration is also dependent on NADH dehydrogenase I. The consequences for energy conservation by anaerobic respiration with NADH as a donor are discussed.Keywordx: NADH dehydrogenase I; fumarate respiration ; proton potential ; menaquinone; Escherichia coli.In respiratory metabolism, Escherichia coli can use a large variety of electron donors and acceptors (Gunsalus, 1992;Unden et al., 1994;Gennis and Stewart, 1996). The conditions resulting in the synthesis of the oxidases and terminal reductases and their physiological roles are largely known (Spiro and Guest, 1991;Gunsalus, 1992;Iuchi and Lin, 1993;Unden et al., 1994. The variety of dehydrogenases that deliver electrons to the quinones is even larger, and for substrates such as formate, H,, glycerol 3-phosphate and NADH, more than one dehydrogenase is found in the bacteria. The isoenzymes catalyze the same reaction (donor + quinone + quinol + oxidized donor), but often they are produced under differing conditions, such as changes in 0, or nitrate supply. NADH is the most important electron donor for the respiratory chains of E. coli during growth with many substrates. E. coli contains two NADH:quinone oxidoreductases (Matsushita et al., 1987;. NADH dehydrogenase I1 is encoded by the ndh gene and is synthesized mainly during aerobic growth due to the transcriptional repression by the 0,-responsive regulator FNR under anoxic conditions (Spiro et al., 1989;Green and Guest, 1994). The alternative enzyme, NADH dehydrogenase I is encoded by the nuo operon and consists of 14 subunits (Weidner et al., 1993). The enzyme is thought to translocate 2 H+/e-coupled to the redox reaction (Friedrich et al., 1995). The nu0 operon is preceded by a large intergenic region of 650 bp and is subject to complex transcriptional regulation electron acceptors 0, and nitrate compared with fermentative growth, in agreement with the significance of the enzyme in respiratory metabolism. The 0,-dependent regulation consisted mainly of repression by ArcA under an...

In an oxystat, the synthesis of the fermentation products formate, acetate, ethanol, lactate, and succinate of Escherichia coli was studied as a function of the O2 tension (pO2) in the medium. The pO2 values that gave rise to half-maximal synthesis of the products (pO0. 5) were 0.2-0.4 mbar for ethanol, acetate, and succinate, and 1 mbar for formate. The pO0.5 for the expression of the adhE gene encoding alcohol dehydrogenase was approximately 0.8 mbar. Thus, the pO2 for the onset of fermentation was distinctly lower than that for anaerobic respiration (pO0.5

The heterofermentative lactic acid bacterium Oenococcus oeni requires pantothenic acid for growth. In the presence of sufficient pantothenic acid, glucose was converted by heterolactic fermentation stoichiometrically to lactate, ethanol and CO2. Under pantothenic acid limitation, substantial amounts of erythritol, acetate and glycerol were produced by growing and resting bacteria. Production of erythritol and glycerol was required to compensate for the decreasing ethanol production and to enable the synthesis of acetate. In ribose fermentation, there were no shifts in the fermentation pattern in response to pantothenate supply. In the presence of pantothenate, growing O. oeni contained at least 10.2 microM HSCoA, whereas the HSCoA content was tenfold lower after growth in pantothenate-depleted media. HSCoA and acetyl-CoA are cosubstrates of phosphotransacetylase and acetaldehyde dehydrogenase from the ethanol pathway. Both enzymes were found with activities commensurate with their function in ethanol production during heterolactic fermentation. From the kinetic data of the enzymes and the HSCoA and acetyl-CoA contents, it can be calculated that, under pantothenate limitation, phosphotransacetylase, and in particular acetaldehyde dehydrogenase activities become limiting due to low levels of the cosubstrates. Thus HSCoA deficiency represents the major limiting factor in heterolactic fermentation of glucose under pantothenate deficiency and the reason for the shift to erythritol, acetate, and glycerol fermentation.

The operation of the citric acid cycle of Escherichia coli during nitrate respiration (anoxic conditions) was studied by measuring end products and enzyme activities. Excretion of products other than CO2, such as acetate or ethanol, was taken as an indication for a non-functional cycle. From glycerol, approximately 0.3 mol acetate was produced; the residual portion was completely oxidized, indicating the presence of a partially active citric acid cycle. In an arcA mutant devoid of the transcriptional regulator ArcA, glycerol was completely oxidized with nitrate as an electron acceptor, demonstrating derepression and function of the complete pathway. Glucose, on the other hand, was excreted mostly as acetate by the wild-type and by the arcA mutant. During growth on glucose, but not on glycerol, activities of succinate dehydrogenase and of 2-oxoglutarate dehydrogenase were missing nearly completely. Thus, the previously described strong repression of the citric acid cycle during nitrate respiration occurs only during growth on glucose and is the effect of anaerobic and, more important, of glucose repression. In Pseudomonas fluorescens (but not Pseudomonas stutzeri), a similar decrease of citric acid cycle function during anaerobic growth with nitrate was found, indicating a broad distribution of this regulatory principle.