Structural and Functional insights into the catalytic mechanism of the Type II NADH:quinone oxidoreductase family

Marreiros, Bruno C.; Sena, Filipa V.; Sousa, Filipe de; Oliveira, A. Sofia F.; Soares, Cláudio M.; Batista, Ana P.; Pereira, Manuela M.

doi:10.1038/srep42303

Cited by 27 publications

(14 citation statements)

References 30 publications

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“…According to the proposed catalytic mechanism of Marreiros et al [22] , upon binding of NADH, FAD is reduced through hydride transfer to the N5 of the FAD isoalloxazine ring. The second proton needed for the full protonation of the reduced FAD is suggested to be provided by rearrangement of the hydrogen bond network around N1 in which a strictly conserved D302 is present.…”

Section: Resultsmentioning

confidence: 99%

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Regulation of the mechanism of Type-II NADH: Quinone oxidoreductase from S. aureus

et al. 2018

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Type-II NADH:quinone oxidoreductases (NDH-2s) are membrane proteins involved in respiratory chains and the only enzymes with NADH:quinone oxidoreductase activity expressed in Staphylococcus aureus (S. aureus), one of the most common causes of clinical infections. NDH-2s are members of the two-Dinucleotide Binding Domains Flavoprotein (tDBDF) superfamily, having a flavin adenine dinucleotide, FAD, as prosthetic group and NAD(P)H as substrate. The establishment of a Charge-Transfer Complex (CTC) between the isoalloxazine ring of the reduced flavin and the nicotinamide ring of NAD+ in NDH-2 was described, and in this work we explored its role in the kinetic mechanism using different electron donors and electron acceptors. We observed that CTC slows down the rate of the second half reaction (quinone reduction) and determines the effect of HQNO, an inhibitor. Also, protonation equilibrium simulations clearly indicate that the protonation probability of an important residue for proton transfer to the active site (D302) is influenced by the presence of the CTC. We propose that CTC is critical for the overall mechanism of NDH-2 and possibly relevant to keep a low quinol/quinone ratio and avoid excessive ROS production in vivo.

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Section: Resultsmentioning

confidence: 99%

“…Marreiros et al [22] , recently proposed the first catalytic mechanism for NDH-2 family. In this study, which contemplates CTC formation coincidentally with NADH binding, aspartate 302 (D302), present in the vicinity of FAD, was observed to be totally conserved (100%), even among other members of the tDBDF superfamily.…”

Section: Introductionmentioning

confidence: 99%

Regulation of the mechanism of Type-II NADH: Quinone oxidoreductase from S. aureus

et al. 2018

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“…The primary structure of NDH-2 commonly includes two GXGXXG motifs within the b-sheet-a-helix-b-sheet domains (Rossman fold), one for binding NAD(P)H and another for FAD or FMN (Wierenga et al, 1986). Although the crystal structures of NDH-2 from Saccharomyces cerevisiae (Ndi1; Feng et al, 2012) and the bacterial type NDH-2 from Caldalkalibacillus thermarum (Heikal et al, 2014) as well as from Staphylococcus aureus (Sena et al, 2015) have been solved, the actual mechanism of NADH:quinone oxidoreduction still remains unclear despite recently presented models (Marreiros et al, 2017). It is generally agreed that, in organisms where NADH oxidation is carried out solely by NDH-2s, the main function of these enzymes is to perform the respiratory chainlinked NADH turnover and the following donation of electrons to the protein complexes that produce the H + gradient powering ATP production (for review, see Melo et al, 2004).…”

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confidence: 99%

Role of Type 2 NAD(P)H Dehydrogenase NdbC in Redox Regulation of Carbon Allocation in Synechocystis

et al. 2017

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H dehydrogenases comprise type 1 (NDH-1) and type 2 (NDH-2s) enzymes. Even though the NDH-1 complex is a wellcharacterized protein complex in the thylakoid membrane of Synechocystis sp. PCC 6803 (hereafter Synechocystis), the exact roles of different NDH-2s remain poorly understood. To elucidate this question, we studied the function of NdbC, one of the three NDH-2s in Synechocystis, by constructing a deletion mutant (DndbC) for a corresponding protein and submitting the mutant to physiological and biochemical characterization as well as to comprehensive proteomics analysis. We demonstrate that the deletion of NdbC, localized to the plasma membrane, affects several metabolic pathways in Synechocystis in autotrophic growth conditions without prominent effects on photosynthesis. Foremost, the deletion of NdbC leads, directly or indirectly, to compromised sugar catabolism, to glycogen accumulation, and to distorted cell division. Deficiencies in several sugar catabolic routes were supported by severe retardation of growth of the DndbC mutant under light-activated heterotrophic growth conditions but not under mixotrophy. Thus, NdbC has a significant function in regulating carbon allocation between storage and the biosynthesis pathways. In addition, the deletion of NdbC increases the amount of cyclic electron transfer, possibly via the NDH-1 2 complex, and decreases the expression of several transporters in ambient CO 2 growth conditions.

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“…The aromatic character at W63 has been reported to be highly conserved among NDH-2s (Fig. 5 ) and suggested to stabilize the prosthetic group FAD in the enzyme 32 . The present study therefore identified a new role in substrate binding for these residues.…”

Section: Discussionmentioning

confidence: 95%

Ubiquinone binding site of yeast NADH dehydrogenase revealed by structures binding novel competitive- and mixed-type inhibitors

Yamashita

Inaoka

Shiba

et al. 2018

Sci Rep

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Yeast Ndi1 is a monotopic alternative NADH dehydrogenase. Its crystal structure in complex with the electron acceptor, ubiquinone, has been determined. However, there has been controversy regarding the ubiquinone binding site. To address these points, we identified the first competitive inhibitor of Ndi1, stigmatellin, along with new mixed-type inhibitors, AC0-12 and myxothiazol, and thereby determined the crystal structures of Ndi1 in complexes with the inhibitors. Two separate binding sites of stigmatellin, STG-1 and STG-2, were observed. The electron density at STG-1, located at the vicinity of the FAD cofactor, further demonstrated two binding modes: STG-1a and STG-1b. AC0-12 and myxothiazol are also located at the vicinity of FAD. The comparison of the binding modes among stigmatellin at STG-1, AC0-12, and myxothiazol revealed a unique position for the aliphatic tail of stigmatellin at STG-1a. Mutations of amino acid residues that interact with this aliphatic tail at STG-1a reduced the affinity of Ndi1 for ubiquinone. In conclusion, the position of the aliphatic tail of stigmatellin at STG-1a provides a structural basis for its competitive inhibition of Ndi1. The inherent binding site of ubiquinone is suggested to overlap with STG-1a that is distinct from the binding site for NADH.

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Structural and Functional insights into the catalytic mechanism of the Type II NADH:quinone oxidoreductase family

Cited by 27 publications

References 30 publications

Regulation of the mechanism of Type-II NADH: Quinone oxidoreductase from S. aureus

Regulation of the mechanism of Type-II NADH: Quinone oxidoreductase from S. aureus

Role of Type 2 NAD(P)H Dehydrogenase NdbC in Redox Regulation of Carbon Allocation in Synechocystis

Ubiquinone binding site of yeast NADH dehydrogenase revealed by structures binding novel competitive- and mixed-type inhibitors

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