Conserved Amino Acid Residues of the NuoD Segment Important for Structure and Function of Escherichia coli NDH-1 (Complex I)

Abstract

The NuoD segment (homologue of mitochondrial 49 kDa subunit) of the proton-translocating NADH:quinone oxidoreductase (complex I/NDH-1) from Escherichia coli is in the hydrophilic domain and bears many highly conserved amino acid residues. The three-dimensional structural model of NDH-1 suggests that the NuoD segment, together with the neighboring subunits, constitutes a putative quinone binding cavity. We used the homologous DNA recombination technique to clarify the role of selected key amino acid residues of the NuoD segment. Among them, residues Tyr273 and His224 were considered candidates for having important interactions with the quinone headgroup. Mutant Y273F retained partial activity but lost sensitivity to capsaicin-40. Mutant H224R scarcely affected the activity, suggesting that this residue may not be essential. His224 is located in a loop near the N-terminus of the NuoD segment (Gly217–Phe227) which is considered to form part of the quinone binding cavity. In contrast to the His224 mutation, mutants G217V, P218A, and G225V almost completely lost the activity. One region of this loop is positioned close to a cytosolic loop of the NuoA subunit in the membrane domain, and together they seem to be important in keeping the quinone binding cavity intact. The structural role of the longest helix in the NuoD segment located behind the quinone binding cavity was also investigated. Possible roles of other highly conserved residues of the NuoD segment are discussed.

A Self‐Assembled Respiratory Chain that Catalyzes NADH Oxidation by Ubiquinone‐10 Cycling between Complex I and the Alternative Oxidase

“…The mechanism of Q 10 reduction, an integral part of the unknown coupling mechanism, presents a much greater challenge owing to the extreme hydrophobicity of Q 10 . Previous studies have addressed the effects of inhibitors2 and mutations in and around the substrate binding site,3, 4 and made use of spectroscopy to search for ubiqsemiquinone intermediates 5. However, they have relied on either Q 10 in native membranes5, 6 (which contain many different enzymes, thus complicating spectroscopic and kinetic analyses) or on non‐physiological hydrophilic Q 10 analogues such as ubiquinone‐1, ubiquinone‐2 and decylubiquinone (DQ)4, 7 (which must be added in excessive concentrations to maintain steady‐state catalysis and that react adventitiously at the flavin to generate damaging reactive oxygen and semiquinone species8).…”

Noisy Response to Antibiotic Stress Predicts Subsequent Single-Cell Survival in an Acidic Environment

“…Interestingly, the DnuoC strain showed an aggravated growth rate drop ( Figure S6B) and prolonged and amplified gadBC expression ( Figure S6A) and pH drop ( Figure S6C). NuoC is an essential component for the proper formation of the proton-pumping NADH dehydrogenase I complex (Sinha et al, 2015), which can likely protect from mild acid stress (Kanjee and Houry, 2013; Krulwich et al, 2011). Other respiratory chain mutants did not show a strongly altered response.…”

Food derived respiratory complex I inhibitors modify the effect of Leber hereditary optic neuropathy mutations

“…Maybe this new amino acid favors a tighter interaction with acetogenins and other compounds, such as capsaicin. The interface between the 49 kDa and p. MT-ND1 subunits forms the inhibitor/Q-site, although it is considered that different CI inhibitors act on the same Q-site with different but partially overlapping sites (Okun et al, 1999;Sinha et al, 2015).…”