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Molecules (Basel, Switzerland) Jul 2023This review uses the National Cancer Institute (NCI) COMPARE program to establish an extensive list of heterocyclic iminoquinones and quinones with similarities in... (Review)
Review
This review uses the National Cancer Institute (NCI) COMPARE program to establish an extensive list of heterocyclic iminoquinones and quinones with similarities in differential growth inhibition patterns across the 60-cell line panel of the NCI Developmental Therapeutics Program (DTP). Many natural products and synthetic analogues are revealed as potential NAD(P)H:quinone oxidoreductase 1 (NQO1) substrates, through correlations to dipyridoimidazo[5,4-]benzimidazoleiminoquinone (DPIQ), and as potential thioredoxin reductase (TrxR) inhibitors, through correlations to benzo[1,2,4]triazin-7-ones and pleurotin. The strong correlation to NQO1 infers the enzyme has a major influence on the amount of the active compound with benzo[]perimidines, phenoxazinones, benz[]pyrido[1,2-]indole-6,11-quinones, seriniquinones, kalasinamide, indolequinones, and furano[2,3-]naphthoquinones, hypothesised as prodrugs. Compounds with very strong correlations to known TrxR inhibitors had inverse correlations to the expression of both reductase enzymes, NQO1 and TrxR, including naphtho[2,3-][1,4]oxazepane-6,11-diones, benzo[]carbazole-1,4-diones, pyranonaphthoquinones (including kalafungin, nanaomycin A, and analogues of griseusin A), and discorhabdin C. Quinoline-5,8-dione scaffolds based on streptonigrin and lavendamycin can correlate to either reductase. Inhibitors of TrxR are not necessarily (imino)quinones, e.g., parthenolides, while oxidising moieties are essential for correlations to NQO1, as with the mitosenes. Herein, an overview of synthetic methods and biological activity of each family of heterocyclic imino(quinone) is provided.
Topics: United States; National Cancer Institute (U.S.); Quinones; Indolequinones; Oxidoreductases; NAD(P)H Dehydrogenase (Quinone); Antineoplastic Agents; Neoplasms
PubMed: 37446864
DOI: 10.3390/molecules28135202 -
Biochemistry Oct 2022The enzyme nicotine oxidoreductase (NicA2) is a member of the flavoprotein amine oxidase family that uses a cytochrome c protein (CycN) as its oxidant instead of...
The enzyme nicotine oxidoreductase (NicA2) is a member of the flavoprotein amine oxidase family that uses a cytochrome c protein (CycN) as its oxidant instead of dioxygen, which is the oxidant used by most other members of this enzyme family. We recently identified a potential binding site for CycN on the surface of NicA2 through rigid body docking [. , (8), 102251]. However, this potential binding interface has not been experimentally validated. In this paper, we used unnatural amino acid incorporation to probe the binding interface between NicA2 and CycN. Our results are consistent with a structural model of the NicA2-CycN complex predicted by protein-protein docking and AlphaFold, suggesting that this is the binding site for CycN on NicA2's surface. Based on additional mutagenesis of potentially redox active residues in NicA2, we propose that electron transfer from NicA2's flavin to CycN's heme occurs without the assistance of a protein-derived wire.
Topics: Amines; Amino Acids; Cytochromes c; Electron Transport; Electrons; Flavins; Flavoproteins; Heme; Nicotine; Oxidants; Oxidation-Reduction; Oxidoreductases; Oxygen
PubMed: 36154019
DOI: 10.1021/acs.biochem.2c00472 -
Current Opinion in Structural Biology Aug 2018Though established 40 years ago, the field of de novo protein design has recently come of age, with new designs exhibiting an unprecedented level of sophistication in... (Review)
Review
Though established 40 years ago, the field of de novo protein design has recently come of age, with new designs exhibiting an unprecedented level of sophistication in structure and function. With respect to catalysis, de novo enzymes promise to revolutionise the industrial production of useful chemicals and materials, while providing new biomolecules as plug-and-play components in the metabolic pathways of living cells. To this end, there are now de novo metalloenzymes that are assembled in vivo, including the recently reported C45 maquette, which can catalyse a variety of substrate oxidations with efficiencies rivalling those of closely related natural enzymes. Here we explore the successful design of this de novo enzyme, which was designed to minimise the undesirable complexity of natural proteins using a minimalistic bottom-up approach.
Topics: Binding Sites; Catalysis; Models, Molecular; Oxidation-Reduction; Oxidoreductases; Protein Binding; Protein Engineering; Protein Interaction Domains and Motifs; Recombinant Proteins; Structure-Activity Relationship; Substrate Specificity
PubMed: 29754103
DOI: 10.1016/j.sbi.2018.04.008 -
The Journal of Biological Chemistry 2021Environmental sequence data of microbial communities now makes up the majority of public genomic information. The assignment of a function to sequences from these...
Environmental sequence data of microbial communities now makes up the majority of public genomic information. The assignment of a function to sequences from these metagenomic sources is challenging because organisms associated with the data are often uncharacterized and not cultivable. To overcome these challenges, we created a rationally designed expression library of metagenomic proteins covering the sequence space of the thioredoxin superfamily. This library of 100 individual proteins represents more than 22,000 thioredoxins found in the Global Ocean Sampling data set. We screened this library for the functional rescue of Escherichia coli mutants lacking the thioredoxin-type reductase (ΔtrxA), isomerase (ΔdsbC), or oxidase (ΔdsbA). We were able to assign functions to more than a quarter of our representative proteins. The in vivo function of a given representative could not be predicted by phylogenetic relation but did correlate with the predicted isoelectric surface potential of the protein. Selected proteins were then purified, and we determined their activity using a standard insulin reduction assay and measured their redox potential. An unexpected gel shift of protein E5 during the redox potential determination revealed a redox cycle distinct from that of typical thioredoxin-superfamily oxidoreductases. Instead of the intramolecular disulfide bond formation typical for thioredoxins, this protein forms an intermolecular disulfide between the attacking cysteines of two separate subunits during its catalytic cycle. Our functional metagenomic approach proved not only useful to assign in vivo functions to representatives of thousands of proteins but also uncovered a novel reaction mechanism in a seemingly well-known protein superfamily.
Topics: Catalysis; Cysteine; Environmental Monitoring; Escherichia coli; Glutaredoxins; Metagenomics; Multigene Family; Oceans and Seas; Oxidation-Reduction; Phylogeny; Protein Disulfide-Isomerases; Thioredoxin-Disulfide Reductase; Thioredoxins
PubMed: 33361108
DOI: 10.1074/jbc.RA120.016350 -
The FEBS Journal Apr 2020Promiscuity is the coincidental ability of an enzyme to catalyze its native reaction and additional reactions that are not biological functions in the same active site.... (Review)
Review
Promiscuity is the coincidental ability of an enzyme to catalyze its native reaction and additional reactions that are not biological functions in the same active site. Promiscuity plays a central role in enzyme evolution and is thus a useful property for protein and metabolic engineering. This review examines enzyme evolution holistically, beginning with evaluating biochemical support for four enzyme evolution models. As expected, there is strong biochemical support for the subfunctionalization and innovation-amplification-divergence models, in which promiscuity is a central feature. In many cases, however, enzyme evolution is more complex than the models indicate, suggesting much is yet to be learned about selective pressures on enzyme function. A complete understanding of enzyme evolution must also explain the ability of metabolic networks to integrate new enzyme activities. Hidden within metabolic networks are underground metabolic pathways constructed from promiscuous activities. We discuss efforts to determine the diversity and pervasiveness of underground metabolism. Remarkably, several studies have discovered that some metabolic defects can be repaired via multiple underground routes. In prokaryotes, metabolic innovation is driven by connecting enzymes acquired by horizontal gene transfer (HGT) into the metabolic network. Thus, we end the review by discussing how the combination of promiscuity and HGT contribute to evolution of metabolism in prokaryotes. Future studies investigating the contribution of promiscuity to enzyme and metabolic evolution will need to integrate deeper probes into the influence of evolution on protein biophysics, enzymology, and metabolism with more complex and realistic evolutionary models. ENZYMES: lactate dehydrogenase (EC 1.1.1.27), malate dehydrogenase (EC 1.1.1.37), OSBS (EC 4.2.1.113), HisA (EC 5.3.1.16), TrpF, PriA (EC 5.3.1.24), R-mandelonitrile lyase (EC 4.1.2.10), Maleylacetate reductase (EC 1.3.1.32).
Topics: Aldehyde-Lyases; Gene Transfer, Horizontal; Oxidoreductases; Oxidoreductases Acting on CH-CH Group Donors; Substrate Specificity
PubMed: 31858709
DOI: 10.1111/febs.15185 -
Molecules (Basel, Switzerland) Dec 2018All eukaryotic molybdenum (Mo) enzymes contain in their active site a Mo Cofactor (Moco), which is formed by a tricyclic pyranopterin with a dithiolene chelating the Mo... (Review)
Review
All eukaryotic molybdenum (Mo) enzymes contain in their active site a Mo Cofactor (Moco), which is formed by a tricyclic pyranopterin with a dithiolene chelating the Mo atom. Here, the eukaryotic Moco biosynthetic pathway and the eukaryotic Moco enzymes are overviewed, including nitrate reductase (NR), sulfite oxidase, xanthine oxidoreductase, aldehyde oxidase, and the last one discovered, the moonlighting enzyme mitochondrial Amidoxime Reducing Component (mARC). The mARC enzymes catalyze the reduction of hydroxylated compounds, mostly N-hydroxylated (NHC), but as well of nitrite to nitric oxide, a second messenger. mARC shows a broad spectrum of NHC as substrates, some are prodrugs containing an amidoxime structure, some are mutagens, such as 6-hydroxylaminepurine and some others, which most probably will be discovered soon. Interestingly, all known mARC need the reducing power supplied by different partners. For the NHC reduction, mARC uses cytochrome b5 and cytochrome b5 reductase, however for the nitrite reduction, plant mARC uses NR. Despite the functional importance of mARC enzymatic reactions, the structural mechanism of its Moco-mediated catalysis is starting to be revealed. We propose and compare the mARC catalytic mechanism of nitrite versus NHC reduction. By using the recently resolved structure of a prokaryotic MOSC enzyme, from the mARC protein family, we have modeled an in silico three-dimensional structure of a eukaryotic homologue.
Topics: Animals; Cardiac Myosins; Coenzymes; Enzymes; Eukaryotic Cells; Mammals; Metabolic Networks and Pathways; Metalloproteins; Molybdenum; Molybdenum Cofactors; Myosin Light Chains; Nitrate Reductase; Nitrites; Oxidoreductases; Pteridines
PubMed: 30545001
DOI: 10.3390/molecules23123287 -
STAR Protocols Mar 2021Lipid peroxidation of polyunsaturated fatty acid (PUFA) phospholipids induces necrotic cell death through compromised cell membrane integrity during ferroptosis. We...
Lipid peroxidation of polyunsaturated fatty acid (PUFA) phospholipids induces necrotic cell death through compromised cell membrane integrity during ferroptosis. We established assays to investigate oxidoreductase-mediated oxidative rupture, specifically via NADPH-cytochrome P450 reductase (POR) and NADH-cytochrome b5 reductase (CYB5R1), of PUFA phospholipids in artificially generated protein-free liposomes. Liposome breakage was detected via Tb liposome release and electron microscopy liposome morphology imaging. This protocol was also applied to other oxidoreductases with analogous functions and investigation of ferroptotic membrane damage in cell-free systems. For complete details on the use and execution of this protocol, please refer to Yan et al. (2020).
Topics: Animals; Cattle; Cytochrome P-450 Enzyme System; Cytochrome-B(5) Reductase; Fatty Acids, Unsaturated; Humans; Liposomes; Oxidation-Reduction
PubMed: 33718888
DOI: 10.1016/j.xpro.2021.100360 -
Molecules (Basel, Switzerland) Dec 2015Computational methods are well-established tools in the drug discovery process and can be employed for a variety of tasks. Common applications include lead... (Review)
Review
Computational methods are well-established tools in the drug discovery process and can be employed for a variety of tasks. Common applications include lead identification and scaffold hopping, as well as lead optimization by structure-activity relationship analysis and selectivity profiling. In addition, compound-target interactions associated with potentially harmful effects can be identified and investigated. This review focuses on pharmacophore-based virtual screening campaigns specifically addressing the target class of hydroxysteroid dehydrogenases. Many members of this enzyme family are associated with specific pathological conditions, and pharmacological modulation of their activity may represent promising therapeutic strategies. On the other hand, unintended interference with their biological functions, e.g., upon inhibition by xenobiotics, can disrupt steroid hormone-mediated effects, thereby contributing to the development and progression of major diseases. Besides a general introduction to pharmacophore modeling and pharmacophore-based virtual screening, exemplary case studies from the field of short-chain dehydrogenase/reductase (SDR) research are presented. These success stories highlight the suitability of pharmacophore modeling for the various application fields and suggest its application also in futures studies.
Topics: Animals; Drug Discovery; Humans; Hydroxysteroid Dehydrogenases; Oxidoreductases; Structure-Activity Relationship
PubMed: 26703541
DOI: 10.3390/molecules201219880 -
Biochemistry. Biokhimiia Dec 2014The respiratory chain in the majority of organisms with aerobic type metabolism features the concomitant existence of the phosphorylating cytochrome pathway and the... (Review)
Review
The respiratory chain in the majority of organisms with aerobic type metabolism features the concomitant existence of the phosphorylating cytochrome pathway and the cyanide- and antimycin A-insensitive oxidative route comprising a so-called alternative oxidase (AOX) as a terminal oxidase. In this review, the history of AOX discovery is described. Considerable evidence is presented that AOX occurs widely in organisms at various levels of organization and is not confined to the plant kingdom. This enzyme has not been found only in Archaea, mammals, some yeasts and protists. Bioinformatics research revealed the sequences characteristic of AOX in representatives of various taxonomic groups. Based on multiple alignments of these sequences, a phylogenetic tree was constructed to infer their possible evolution. The ways of AOX activation, as well as regulatory interactions between AOX and the main respiratory chain are described. Data are summarized concerning the properties of AOX and the AOX-encoding genes whose expression is either constitutive or induced by various factors. Information is presented on the structure of AOX, its active center, and the ubiquinone-binding site. The principal functions of AOX are analyzed, including the cases of cell survival, optimization of respiratory metabolism, protection against excess of reactive oxygen species, and adaptation to variable nutrition sources and to biotic and abiotic stress factors. It is emphasized that different AOX functions complement each other in many instances and are not mutually exclusive. Examples are given to demonstrate that AOX is an important tool to overcome the adverse aftereffects of restricted activity of the main respiratory chain in cells and whole animals. This is the first comprehensive review on alternative oxidases of various organisms ranging from yeasts and protists to vascular plants.
Topics: Bacteria; Electron Transport; Mitochondrial Proteins; Nutritional Physiological Phenomena; Oxidoreductases; Phylogeny; Plant Proteins; Protein Conformation
PubMed: 25749168
DOI: 10.1134/S0006297914130112 -
Cell Cycle (Georgetown, Tex.) 2017
Topics: Biological Transport; Cell Line, Tumor; Glutathione; Oxidation-Reduction; Oxidoreductases
PubMed: 28933594
DOI: 10.1080/15384101.2017.1377031