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Structural Insights into the NAD(P)H:Quinone Oxidoreductase from Phytophthora capsici.

ABSTRACT: Soluble quinone oxidoreductases catalyze transfer of electrons from NADPH to quinones. Transfer of electrons is essential for detoxification of synthetic compounds. Here, we present the crystal structure of a NADPH-dependent QOR from Phytophthora capsici (Pc) complexed with NADPH at 2.4 Å resolution. The enzyme exhibits a bi-modular architecture, containing a NADPH-binding groove and a substrate-binding pocket in each subunit. In the crystal, each asymmetric unit of PcQOR contains two molecules stabilized by intermolecular interactions. Gel filtration and ultracentrifugation analyses reveal that it functions as a tetramer in solution. Alignment of homologous structures exhibits a conserved topology. However, the active sites vary among the homologues, indicating differences in substrate specificities. Enzymatic assays indicate that PcQOR tends to catalyze the large substrates, like 9,10-phenanthrenequinone. Computational simulation associated with site-directed mutagenesis and enzymatic activity analysis declares a potential quinone-binding channel. The ability to reduce quinones probably helps P. capsici to detoxify some harmful chemicals encountered during invasion.

SUBMITTER: Yang C

PROVIDER: S-EPMC9330140 | biostudies-literature | 2022 Jul

REPOSITORIES: biostudies-literature

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Structural Insights into the NAD(P)H:Quinone Oxidoreductase from Phytophthora capsici.

Yang Cancan C Huang Zhenling Z Zhang Xiuguo X Zhu Chunyuan C

ACS omega 20220713 29

Soluble quinone oxidoreductases catalyze transfer of electrons from NADPH to quinones. Transfer of electrons is essential for detoxification of synthetic compounds. Here, we present the crystal structure of a NADPH-dependent QOR from Phytophthora capsici (Pc) complexed with NADPH at 2.4 Å resolution. The enzyme exhibits a bi-modular architecture, containing a NADPH-binding groove and a substrate-binding pocket in each subunit. In the crystal, each asymmetric unit of PcQOR co ...[more]

PMID: 35910145

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Project description:NAD(P)H:quinone oxidoreductase type I (NQO1) is a target enzyme for triggered delivery of drugs at inflamed tissue and tumor sites, particularly those that challenge traditional therapies. Prodrugs, macromolecules, and molecular assemblies possessing trigger groups that can be cleaved by environmental stimuli are vehicles with the potential to yield active drug only at prescribed sites. Furthermore, quinone propionic acids (QPAs) covalently attached to prodrugs or liposome surfaces can be removed by application of a reductive trigger stimulus, such as that from NQO1; their rates of reductive activation should be tunable via QPA structure. We explored in detail the recombinant human NAD(P)H:quinone oxidoreductase type I (rhNQO1)-catalyzed NADH reduction of a family of substituted QPAs and obtained high precision kinetic parameters. It is found that small changes in QPA structure-in particular, single atom and function group substitutions on the quinone ring at R(1)-lead to significant impacts on the Michaelis constant (K(m)), maximum velocity (V(max)), catalytic constant (k(cat)), and catalytic efficiency (k(cat)/K(m)). Molecular docking simulations demonstrate that alterations in QPA structure result in large changes in QPA alignment and placement with respect to the flavin isoalloxazine ring in the active site of rhNQO1; a qualitative relationship exists between the kinetic parameters and the depth of QPA penetration into the rhNQO1 active site. From a quantitative perspective, a very good correlation is observed between log(k(cat)/K(m)) and the molecular-docking-derived distance between the flavin hydride donor site and quinone hydride acceptor site in the QPAs, an observation that is in agreement with developing theories. The comprehensive kinetic and molecular modeling knowledge obtained for the interaction of recombinant human NQO1 with the quinone propionic acid analogues provides insight into the design and implementation of the QPA trigger groups for drug delivery applications.

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Structural Insights into the NAD(P)H:Quinone Oxidoreductase from Phytophthora capsici.

Publications

Structural Insights into the NAD(P)H:Quinone Oxidoreductase from <i>Phytophthora capsici</i>.

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