Roles of NapF, NapG and NapH, subunits of the Escherichia coli periplasmic nitrate reductase, in ubiquinol oxidation

Abstract

Roles of NapF, NapG and NapH, subunits of the Escherichia coli periplasmic nitrate reductase, in ubiquinol oxidation that NapG and H, but not NapF, are essential for electron transfer from ubiquinol to NapAB. The decreased yield of biomass resulting from loss of NapF in a Ubi + Men+ strain implicates NapF in an energyconserving role coupled to the oxidation of ubiquinol. We propose that NapG and H form an energyconserving quinol dehydrogenase functioning as either components of a proton pump or in a Q cycle, as electrons are transferred from ubiquinol to NapC. IntroductionEnergy-conserving electron transfer pathways in enteric bacteria are usually depicted as a series of substratespecific dehydrogenases feeding electrons into a common quinone pool, from which they are transferred via specific quinol dehydrogenases to cytochrome oxidases during aerobic growth or terminal reductases during anaerobic growth. This is clearly an oversimplification, however, because there are three types of functional quinone in Escherichia coli, ubiquinone 8 (UQ) and the naphthoquinones demethylmenaquinone (DMK) and menaquinone (MK). UQ is generally regarded as the 'aerobic' quinone in the sense that ubiquinone is far more abundant than MK and DMK during aerobic growth (Wallace and Young, 1977;Wissenbach et al., 1992;Soballe and Poole, 1999). Furthermore, UQ is essential for succinoxidase activity. Conversely, the naphthoquinone pool is essential for anaerobic respiration using nitrite, fumarate, dimethyl sulphoxide (DMSO) or trimethylamine N-oxide (TMAO) (Wissenbach et al., 1990;1992;Tyson et al., 1997). The selectivity of quinones for specific electron donors or acceptors can be explained by the difference in mid-point redox potential between the UQ/UQH 2 couple (E m,7 = +113 mV) and the MK/MKH 2 couple (E m,7 = -74 mV) (Soballe and Poole, 1999). In addition, there may also be structural constraints that limit the enzymes of the respiratory chain to binding a specific quinone.Nitrate respiration in E. coli has a unique position as electrons from both UQH 2 and MKH 2 , but not DMKH 2 , can be used for nitrate reduction (Wissenbach et al., 1990;1992;Tyson et al., 1997). E. coli expresses three nitrate reductases. Two of them, nitrate reductases A and Z, are membrane bound and reduce nitrate in the cytoplasm. SummaryThe nap operon of Escherichia coli K-12, encoding a periplasmic nitrate reductase (Nap), encodes seven proteins. The catalytic complex in the periplasm, NapA-NapB, is assumed to receive electrons from the quinol pool via the membrane-bound cytochrome NapC. Like NapA, B and C, a fourth polypeptide, NapD, is also essential for Nap activity. However, none of the remaining three polypeptides, NapF, G and H, which are predicted to encode non-haem, iron-sulphur proteins, are essential for Nap activity, and their function is currently unknown. The relative rates of growth and electron transfer from physiological substrates to Nap have been investigated using strains defective in the two membrane-bound nitrate reductases, and als...

Characterization of the expression and activity of the periplasmic nitrate reductase of Paracoccus pantotrophus in chemostat cultures

“…For example, Nap plays a role in anaerobic denitrification in Pseudomonas G-179 and Rhodobacter sphaeroides f. sp. denitrificans, in anaerobic nitrate respiration under nitrate-limited conditions in E. coli, and redox-poising of the photosynthetic electrontransport chain or survival during light-dark transitions in Rhodobacter capsulatus (Jones et al, 1990;Siddiqui et al, 1993;Castillo et al, 1996;Zumft, 1997;Liu et al, 1999;Moreno-Vivian et al, 1999;Philippot & Hojberg, 1999;Potter et al, 1999Potter et al, , 2001Richardson et al, 2001;Brondijk et al, 2002;Ellington et al, 2003). Previously, the importance of transcriptional regulation for the regulation of Nap activity in P. pantotrophus has been established in batch culture (Ellington et al, 2002).…”

The periplasmic nitrate reductase in Shewanella: the resolution, distribution and functional implications of two NAP isoforms, NapEDABC and NapDAGHB

“…Studies of the oxygen-dependent regulation of quinone biosynthesis in Shewanella show that most, but not all, species produce ubiquinones and menaquinones. The available quinone pool may in turn direct the regulation of the electron transfer from quinol to the NAP isoforms (Brondijk et al, 2002(Brondijk et al, , 2004Potter et al, 2001;Potter & Cole, 1999). The transcriptional regulators that mediate global gene expression between anaerobic and aerobic growth might also play a role in the regulation of the expression of the NAP isoforms in Shewanella.…”

“…Anaerobic cultures of S. oneidensis MR-1 produce four to fivefold more menaquinols than ubiquinols (Myers & Myers, 2000). In E. coli, it has been established that irrespective of the quinol pool (ubiquinol or menaquinol), NapC is essential for NAP activity (Brondijk et al, 2002;Potter et al, 2001;Potter & Cole, 1999). The role of the NapGH complex was found to be dependent upon the quinol source (Brondijk et al, 2002(Brondijk et al, , 2004.…”

“…In E. coli, it has been established that irrespective of the quinol pool (ubiquinol or menaquinol), NapC is essential for NAP activity (Brondijk et al, 2002;Potter et al, 2001;Potter & Cole, 1999). The role of the NapGH complex was found to be dependent upon the quinol source (Brondijk et al, 2002(Brondijk et al, , 2004. Where ubiquinol was the sole quinol source, NapGH was essential as an ubiquinol dehydrogenase that mediated electron transfer between NapC and NapAB; this NapGH-based mediation was not essential in E. coli constructs where menaquinol and ubiquinol were available (Brondijk et al, 2002(Brondijk et al, , 2004.…”

Physiological and evolutionary studies of NAP systems in Shewanella piezotolerans WP3

“…In the napC free strains, such as Campylobacter jejuni and Wolinella succinogenes, NapGH are at the substitution place of NapC (Pittman et al, 2007, Kern andSimon, 2008). In E. coli, which contains NapGH and NapC, NapGH is not involved in menaquinol oxidation and instead form a quinol dehydrogenase that transfers electrons from ubiquinol through NapC to NapAB (Brondijk et al, 2002(Brondijk et al, , 2004. In the well-studied S. oneidensis MR-1, which also doesn't have a napC gene, CymA has key roles in both nitrate and nitrite reduction (Murphy and Saltikov, 2007;Gao et al, 2009).…”