The C-terminal Domain of 4-Hydroxyphenylacetate 3-Hydroxylase from Acinetobacter baumannii Is an Autoinhibitory Domain

Phongsak, Thanawat; Sucharitakul, Jeerus; Thotsaporn, Kittisak; Oonanant, Worrapoj; Yuvaniyama, Jirundon; Svasti, Jisnuson; Ballou, David P.; Chaiyen, Pimchai

doi:10.1074/jbc.m112.354472

Cited by 23 publications

(31 citation statements)

References 48 publications

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“…S10 indicate that FADH Ϫ generated by HadX can be transferred to a monooxygenase without requiring any specific proteinprotein interaction and HadX can also supply FADH Ϫ to other enzyme systems such as to C 2 monooxygenase. Features of the FADH Ϫ free diffusion transfer found in HadX-HadA system are similar to those of the C 1 -C 2 , LuxG-LuxAB, ActVB-ActVA, and HpaC-HpaB systems (20,34,38,46,65).…”

Section: Catalytic Mechanisms Of Hadx Reductase and Hadb Reductasesupporting

confidence: 58%

“…HadX is the first flavin reductase in the degradation pathway of HPs and NPs that has been characterized in-depth. The functional and catalytic properties of HadX and other known flavin reductases including TftC from Burkholderia cepacia (9,30), TcpX from R. eutropha (29), C 1 from A. baumannii (19,20,46), ActVB from S. coelicolor (33)(34)(35), LuxG from P. leiognathi TH1 (36 -38), HpaC from E. coli (65), PheA2 from Bacillus thermoglucosidasius A7 (66), and StyB from Pseudomanas sp. (67) are compared in terms of their general properties and kinetic parameters in Table 2.…”

Section: Catalytic Mechanisms Of Hadx Reductase and Hadb Reductasementioning

confidence: 99%

“…The rate of flavin reduction in C 1 reductase is quite slow in the absence of HPA (k red of 12 s Ϫ1 ). This is due to an autoinhibitory function of a MarR-like regulatory domain (20). Therefore, HadX should be an efficient reductase for supplying FADH Ϫ to any enzymatic systems that require the use of FADH Ϫ .…”

Section: Catalytic Mechanisms Of Hadx Reductase and Hadb Reductasementioning

confidence: 99%

“…The resulting reduced flavin generally does not bind tightly to the reductase, and thus dissociates from the enzyme to bind to and act as a substrate for the monooxygenase (18 -20). Previously, the reductase component of the p-hydroxyphenylacetate hydroxylase system (C 1 ) (19,20) was used as a flavin reductase to supply FADH Ϫ for HadA (16,17). The partner reductase that was co-evolved with HadA for catalyzing dehalogenation and denitration with HadA is currently unknown (Fig.…”

mentioning

confidence: 99%

“…Mechanistic investigation of these enzymes has not been carried out. HadX was also classified in the same group as various reductases of two-component flavin-dependent monooxygenases, such as ActVB reductase from Streptomyces coelicolor (24% identity) (31)(32)(33)(34)(35), RebF from Lechevalieria aerocolonigenes (29% identity) (36), C 1 from Acinetobacter baumannii (19,20) (16% identity), and LuxG from Photobacterium leiognathi TH1 (9% identity) (37)(38)(39). Previous investigations have shown that these reductases can generate reduced flavin and pass it to their oxygenases without a requirement of protein-protein interaction (21).…”

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confidence: 99%

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A complete bioconversion cascade for dehalogenation and denitration by bacterial flavin–dependent enzymes

Pimviriyakul

Chaiyen

2018

Journal of Biological Chemistry

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Edited by F. Peter GuengerichHalogenated phenol and nitrophenols are toxic compounds that are widely accumulated in the environment. Enzymes in the had operon from the bacterium Ralstonia pickettii DTP0602 have the potential for application as biocatalysts in the degradation of many of these toxic chemicals. HadA monooxygenase previously was identified as a two-component reduced FAD (FADH ؊ )-utilizing monooxygenase with dual activities of dehalogenation and denitration. However, the partner enzymes of HadA, that is, the flavin reductase and quinone reductase that provide the FADH ؊ for HadA and reduce quinone to hydroquinone, remain to be identified. In this report, we overexpressed and purified the flavin reductases, HadB and HadX, to investigate their functional and catalytic properties. Our results indicated that HadB is an FMN-dependent quinone reductase that converts the quinone products from HadA to hydroquinone compounds that are more stable and can be assimilated by downstream enzymes in the pathway. Transient kinetics indicated that HadB prefers NADH and menadione as the electron donor and acceptor, respectively. We found that HadX is an FAD-bound flavin reductase, which can generate FADH ؊ for HadA to catalyze dehalogenation or denitration reactions. Thermodynamic and transient kinetic experiments revealed that HadX prefers to bind FAD over FADH؊ and that HadX can transfer FADH ؊ from HadX to HadA via free diffusion. Moreover, HadX rapidly catalyzed NADH-mediated reduction of flavin and provided the FADH ؊ for a monooxygenase of a different system. Combination of all three flavin-dependent enzymes, i.e. HadA/HadB/HadX, reconstituted an effective dehalogenation and denitration cascade, which may be useful for future bioremediation applications. 2 The abbreviations used are: HPs, halogenated phenol; HadA, two-component FADH Ϫ utilizing monooxygenase; HadB, a quinone reductase encoded in the had operon; HadX, a flavin reductase encoded in the had operon; NPs, nitrophenols; FADH Ϫ , reduced flavin adenine dinucleotide; FMNH Ϫ , reduced flavin mononucleotide; 4-NP, 4-nitrophenol; 4-CP, 4-chlorophenol; HQ, hydroquinone; BQ, benzoquinone; C 1 , the flavin reductase component of p-hydroxyphenylacetate 3-hydroxylase from A. buamannii; C 2 , the oxygenase component of p-hydroxyphenylacetate 3-hydroxylase from A.

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Section: Catalytic Mechanisms Of Hadx Reductase and Hadb Reductasesupporting

confidence: 58%

Section: Catalytic Mechanisms Of Hadx Reductase and Hadb Reductasementioning

confidence: 99%

Section: Catalytic Mechanisms Of Hadx Reductase and Hadb Reductasementioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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A complete bioconversion cascade for dehalogenation and denitration by bacterial flavin–dependent enzymes

Pimviriyakul

Chaiyen

2018

Journal of Biological Chemistry

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Creating Flavin Reductase Variants with Thermostable and Solvent‐Tolerant Properties by Rational‐Design Engineering

et al. 2020

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Scheme1.The C 1 -catalyzed reaction thatg enerates FMNH À for monooxygenase-catalyzed reactions.The NADH-regenerating systemm ightb e, for example, glucose/glucose dehydrogenase, glucose 6-phosphate/glucose 6phosphate dehydrogenase, [41,42] or formate/formate dehydrogenase. [43,44] The monooxygenases mightb e, for example, C 2 or bacterialluciferase. Figure 6. Distances between A58/P58 and I14:A )int he wild type, and B) in the A58P variant over 6nsM Ds imulation,a ttemperatures varying from 300-500 K. The interactions between A58/P58 of C) the wild type, and D) the A58P variant and other residues after 6-ns MD simulations.

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Monooxygenation of aromatic compounds by flavin‐dependent monooxygenases

2018

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Many flavoenzymes catalyze hydroxylation of aromatic compounds especially phenolic compounds have been isolated and characterized. These enzymes can be classified as either single-component or two-component flavin-dependent hydroxylases (monooxygenases). The hydroxylation reactions catalyzed by the enzymes in this group are useful for modifying the biological properties of phenolic compounds. This review aims to provide an in-depth discussion of the current mechanistic understanding of representative flavin-dependent monooxygenases including 3-hydroxy-benzoate 4-hydroxylase (PHBH, a single-component hydroxylase), 3-hydroxyphenylacetate 4-hydroxylase (HPAH, a two-component hydroxylase), and other monooxygenases which catalyze reactions in addition to hydroxylation, including 2-methyl-3-hydroxypyridine-5-carboxylate oxygenase (MHPCO, a single-component enzyme that catalyzes aromatic-ring cleavage), and HadA monooxygenase (a two-component enzyme that catalyzes additional group elimination reaction). These enzymes have different unique structural features which dictate their reactivity toward various substrates and influence their ability to stabilize flavin intermediates such as C4a-hydroperoxyflavin. Understanding the key catalytic residues and the active site environments important for governing enzyme reactivity will undoubtedly facilitate future work in enzyme engineering or enzyme redesign for the development of biocatalytic methods for the synthesis of valuable compounds.

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The C-terminal Domain of 4-Hydroxyphenylacetate 3-Hydroxylase from Acinetobacter baumannii Is an Autoinhibitory Domain

Cited by 23 publications

References 48 publications

A complete bioconversion cascade for dehalogenation and denitration by bacterial flavin–dependent enzymes

A complete bioconversion cascade for dehalogenation and denitration by bacterial flavin–dependent enzymes

Creating Flavin Reductase Variants with Thermostable and Solvent‐Tolerant Properties by Rational‐Design Engineering

Monooxygenation of aromatic compounds by flavin‐dependent monooxygenases

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