Flavin dependent monooxygenases

Huijbers, Mieke M.E.; Montersino, S.; Westphal, Adrie H.; Tischler, Dirk; Berkel, Willem J.H. van

doi:10.1016/j.abb.2013.12.005

Cited by 450 publications

(540 citation statements)

References 119 publications

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“…Slow oxidase activity has been observed in the case of NADH:quinone oxidoreductases (27), chromate reductase (28), flavin-dependent monooxygenases (uncoupling) (16), and old yellow enzyme (29,30). In contrast to PA1024, FAD-dependent NAD(P)H:quinone oxidoreductase 1 (NQO1) and the quinone reductase Lot6p are not oxidized by molecular oxygen (25).…”

Section: Exx(t//s)xlhxrxhxn(t/s)xr(v/i)mentioning

confidence: 99%

Functional Annotation of a Presumed Nitronate Monoxygenase Reveals a New Class of NADH:Quinone Reductases

Ball¹,

Salvi²,

Gadda³

2016

Journal of Biological Chemistry

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The protein PA1024 from Pseudomonas aeruginosa PAO1 is currently classified as 2-nitropropane dioxygenase, the previous name for nitronate monooxygenase in the GenBank TM and PDB databases, but the enzyme was not kinetically characterized. In this study, PA1024 was purified to high levels, and the enzymatic activity was investigated by spectroscopic and polarographic techniques. Purified PA1024 did not exhibit nitronate monooxygenase activity; however, it displayed NADH:quinone reductase and a small NADH:oxidase activity. The enzyme preferred NADH to NADPH as a reducing substrate. PA1024 could reduce a broad spectrum of quinone substrates via a Ping Pong Bi Bi steady-state kinetic mechanism, generating the corresponding hydroquinones. The reductive half-reaction with NADH showed a k red value of 24 s ؊1 and an apparent K d value estimated in the low micromolar range. The enzyme was not able to reduce the azo dye methyl red, routinely used in the kinetic characterization of azoreductases. Finally, we revisited and modified the existing six conserved motifs of PA1024, which define a new class of NADH:quinone reductases and are present in more than 490 hypothetical proteins in the GenBank TM , the vast majority of which are currently misannotated as nitronate monooxygenase.

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Section: Exx(t//s)xlhxrxhxn(t/s)xr(v/i)mentioning

confidence: 99%

Functional Annotation of a Presumed Nitronate Monoxygenase Reveals a New Class of NADH:Quinone Reductases

Ball¹,

Salvi²,

Gadda³

2016

Journal of Biological Chemistry

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“…Then, the flavin derivative performs a nucleophilic or an electrophilic attack, depending on the protonation state of the flavin derivative, which leads to the oxygenation of the substrate, the production of one water molecule and the formation of the oxidized state of the flavin. The role of the NAD(P)H is to restore the reduced form of the flavin in the catalytic site [37,140].…”

Section: Ec 113 and 114: Oxygenases And Monooxygenasesmentioning

confidence: 99%

Review of NAD(P)H-dependent oxidoreductases: Properties, engineering and application

Vidal

Kelly

Mordaka

et al. 2018

Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics

248

147

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A B S T R A C T NAD(P)H-dependent oxidoreductases catalyze the reduction or oxidation of a substrate coupled to the oxidation or reduction, respectively, of a nicotinamide adenine dinucleotide cofactor NAD(P)H or NAD(P) + . NAD(P)Hdependent oxidoreductases catalyze a large variety of reactions and play a pivotal role in many central metabolic pathways. Due to the high activity, regiospecificity and stereospecificity with which they catalyze redox reactions, they have been used as key components in a wide range of applications, including substrate utilization, the synthesis of chemicals, biodegradation and detoxification. There is great interest in tailoring NAD(P)H-dependent oxidoreductases to make them more suitable for particular applications. Here, we review the main properties and classes of NAD(P)H-dependent oxidoreductases, the types of reactions they catalyze, some of the main protein engineering techniques used to modify their properties and some interesting examples of their modification and application.

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“…A reductase component of these enzymes catalyzes flavin reduction, and an oxygenase component catalyzes substrate oxygenation (22,23). The most well studied enzyme in this family is p-hydroxyphenylacetate hydroxylase (HPAH) from Acinetobacter baumannii.…”

mentioning

confidence: 99%

“…Based on sequence similarity, HadA belongs to the family of two-component flavin-dependent monooxygenases that utilize two separate proteins in their catalysis (class D monooxygenase) (22). A reductase component of these enzymes catalyzes flavin reduction, and an oxygenase component catalyzes substrate oxygenation (22,23).…”

mentioning

confidence: 99%

Kinetic Mechanism of the Dechlorinating Flavin-dependent Monooxygenase HadA

Pimviriyakul

Thotsaporn

Sucharitakul

et al. 2017

Journal of Biological Chemistry

108

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Edited by F. Peter GuengerichThe accumulation of chlorophenols (CPs) in the environment, due to their wide use as agrochemicals, has become a serious environmental problem. These organic halides can be degraded by aerobic microorganisms, where the initial steps of various biodegradation pathways include an oxidative dechlorinating process in which chloride is replaced by a hydroxyl substituent. Harnessing these dechlorinating processes could provide an opportunity for environmental remediation, but detailed catalytic mechanisms for these enzymes are not yet known. To close this gap, we now report transient kinetics and product analysis of the dechlorinating flavin-dependent monooxygenase, HadA, from the aerobic organism Ralstonia pickettii DTP0602, identifying several mechanistic properties that differ from other enzymes in the same class. We first overexpressed and purified HadA to homogeneity. Analyses of the products from single and multiple turnover reactions demonstrated that HadA prefers 4-CP and 2-CP over CPs with multiple substituents. Stopped-flow and rapid-quench flow experiments of HadA with 4-CP show the involvement of specific intermediates (C4a-hydroperoxy-FAD and C4a-hydroxy-FAD) in the reaction, define rate constants and the order of substrate binding, and demonstrate that the hydroxylation step occurs prior to chloride elimination. The data also identify the non-productive and productive paths of the HadA reactions and demonstrate that product formation is the rate-limiting step. This is the first elucidation of the kinetic mechanism of a two-component flavin-dependent monooxygenase that can catalyze oxidative dechlorination of various CPs, and as such it will serve as the basis for future investigation of enzyme variants that will be useful for applications in detoxifying chemicals hazardous to human health.

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Flavin dependent monooxygenases

Cited by 450 publications

References 119 publications

Functional Annotation of a Presumed Nitronate Monoxygenase Reveals a New Class of NADH:Quinone Reductases

Functional Annotation of a Presumed Nitronate Monoxygenase Reveals a New Class of NADH:Quinone Reductases

Review of NAD(P)H-dependent oxidoreductases: Properties, engineering and application

Kinetic Mechanism of the Dechlorinating Flavin-dependent Monooxygenase HadA

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