The styrene monooxygenase system

Gassner, George

doi:10.1016/bs.mie.2019.03.019

Cited by 12 publications

(10 citation statements)

References 18 publications

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“…It can be estimated that GnIndA has an uncoupling rate of 86% while GrStyA shows 91% uncoupling. This uncoupling effect was also described for other enzymes, is typically influenced by the type of electron donor, and was reported for NAD(P)H-mimics before [31][32][33]. Nevertheless, this obvious drawback of BNAH can be overcompensated by continuous addition of the cosubstrate ( Figure S3) which is justified by the low price and easy provision of the compound.…”

Section: Substrate Spectrumsupporting

confidence: 63%

Enantioselective Epoxidation by Flavoprotein Monooxygenases Supported by Organic Solvents

et al. 2020

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Styrene and indole monooxygenases (SMO and IMO) are two-component flavoprotein monooxygenases composed of a nicotinamide adenine dinucleotide (NADH)-dependent flavin adenine dinucleotide (FAD)-reductase (StyB or IndB) and a monooxygenase (StyA or IndA). The latter uses reduced FAD to activate oxygen and to oxygenate the substrate while releasing water. We circumvented the need for the reductase by direct FAD reduction in solution using the NAD(P)H-mimic 1-benzyl-1,4-dihydronicotinamide (BNAH) to fuel monooxygenases without NADH requirement. Herein, we report on the hitherto unknown solvent tolerance for the indole monooxygenase from Gemmobacter nectariphilus DSM15620 (GnIndA) and the styrene monooxygenase from Gordonia rubripertincta CWB2 (GrStyA). These enzymes were shown to convert bulky and rather hydrophobic styrene derivatives in the presence of organic cosolvents. Subsequently, BNAH-driven biotransformation was furthermore optimized with regard to the applied cosolvent and its concentration as well as FAD and BNAH concentration. We herein demonstrate that GnIndA and GrStyA enable selective epoxidations of allylic double bonds (up to 217 mU mg−1) in the presence of organic solvents such as tetrahydrofuran, acetonitrile, or several alcohols. Notably, GnIndA was found to resist methanol concentrations up to 25 vol.%. Furthermore, a diverse substrate preference was determined for both enzymes, making their distinct use very interesting. In general, our results seem representative for many IMOs as was corroborated by in silico mutagenetic studies.

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Section: Substrate Spectrumsupporting

confidence: 63%

Enantioselective Epoxidation by Flavoprotein Monooxygenases Supported by Organic Solvents

et al. 2020

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“…LQ26 [67], Pseudomonas sp. VLB120 [50], and P. putida S12 [68]. It may be suggested that bacteria represented in our libraries by the OTUs with the highest similarity of their 16S rRNA gene sequences (>99-100%) to those of styrene-degrading reference bacteria (Appendix A, Table A2, Figure A7) are the most probable potential styrene degraders.…”

Section: Discussionmentioning

confidence: 95%

Biodiversity of Microorganisms Colonizing the Surface of Polystyrene Samples Exposed to Different Aqueous Environments

et al. 2020

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The contamination of marine and freshwater ecosystems with the items from thermoplastics, including polystyrene (PS), necessitates the search for efficient microbial degraders of these polymers. In the present study, the composition of prokaryotes in biofilms formed on PS samples incubated in seawater and the industrial water of a petrochemical plant were investigated. Using a high-throughput sequencing of the V3–V4 region of the 16S rRNA gene, the predominance of Alphaproteobacteria (Blastomonas), Bacteroidetes (Chryseolinea), and Gammaproteobacteria (Arenimonas and Pseudomonas) in the biofilms on PS samples exposed to industrial water was revealed. Alphaproteobacteria (Erythrobacter) predominated on seawater-incubated PS samples. The local degradation of the PS samples was confirmed by scanning microscopy. The PS-colonizing microbial communities in industrial water differed significantly from the PS communities in seawater. Both communities have a high potential ability to carry out the carbohydrates and amino acids metabolism, but the potential for xenobiotic degradation, including styrene degradation, was relatively higher in the biofilms in industrial water. Bacteria of the genera Erythrobacter, Maribacter, and Mycobacterium were potential styrene-degraders in seawater, and Pseudomonas and Arenimonas in industrial water. Our results suggest that marine and industrial waters contain microbial populations potentially capable of degrading PS, and these populations may be used for the isolation of efficient PS degraders.

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“…To determine the effect of the protein environment on the Mn(III/II) reduction potential, spectrophotometric dithionite titration experiments were performed (see the Supporting Information). The E 1/2 (Mn III/II ) for holo-MPP1 was found to be −107 (±5) mV vs SHE (pH 7, Figure S8). ,, Because redox-coupled protonation/deprotonation play critical roles in catalysis, it was important to assess changes in the protonation state accompanying changes in the oxidation state of the Mn ion.…”

Section: Resultsmentioning

confidence: 99%

“…Equilibrium spectrochemical redox titrations. 7 To a 2 mL solution of 15µM Mn(III)MPP1 in 50 mM phosphate, 150 mM NaCl pH 7 buffer, a redox indicator dye (Indigo-5-5'-disulfonic acid, IDIS, with midpoint reduction potential at neutral pH, Em,7 = −125 mV versus SHE) was added to a 28 μM concentration. The sample was then transferred to an anaerobic quartz cuvette and degassed by sequential vacuum pumping and purging with N2.…”

Section: Circular Dichroism (Cd)mentioning

confidence: 99%

“…On the other hand, synthetic chemists have developed a massive library of metalloporphyrins to recapitulate the reactions performed in nature (albeit often less selectively and at slower rates) but also to perform reactions not found in Nature: such as cyclopropanation and amination. 2,[4][5] While directed evolution has produced engineered heme proteins with impressive rates and selectivities for abiological reactions [6][7] , de novo protein design offers the opportunity to test our understanding of protein structure-function relationships. [8][9][10][11][12] A complete understanding of how proteins tune the function of heme cofactors combined with novel reactivity of synthetic cofactors, would allow for the design of functional metalloproteins that can expand upon the scope of natural proteins.…”

Section: Introductionmentioning

confidence: 99%

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De Novo Design, Solution Characterization, and Crystallographic Structure of an Abiological Mn–Porphyrin-Binding Protein Capable of Stabilizing a Mn(V) Species

et al. 2020

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De novo protein design offers the opportunity to test our understanding of how metalloproteins perform difficult transformations. Attaining high-resolution structural information is critical to understanding how such designs function. There have been many successes in the design of porphyrin-binding proteins, however crystallographic characterization has been elusive, limiting what can be learned from such studies as well as the extension to new functions. Moreover, formation of highly oxidizing high-valent intermediates poses design challenges that have not been previously implemented: 1) purposeful design of substrate/oxidant access to the binding site and 2) limiting deleterious oxidation of the protein scaffold. Here we report the first crystallographically characterized porphyrin-binding protein that was programmed to not only bind a synthetic Mn-porphyrin but also maintain binding site access to form high-valent oxidation states. We explicitly designed a binding site with accessibility to dioxygen units in the open coordination site of the Mn center. In solution, the protein is capable of accessing a high-valent Mn(V)-oxo species which can transfer an O-atom to a thioether substrate. The crystallographic structure is within 0.6 Å of the design, and indeed contained an aquo ligand with a second water molecule stabilized by hydrogen-bonding to a Gln sidechain in the active site, offering a structural explanation for the observed reactivity. File list (2) download file view on ChemRxiv MnDPP_manuscript_092820_FINAL_chemrxiv.pdf (6.62 MiB) download file view on ChemRxiv MnDPP_SI_092820_FINAL.pdf (4.08 MiB) De novo design, solution characterization, and crystallographic structure of an abiological Mn-porphyrin binding protein capable of stabilizing a Mn(V) species

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The styrene monooxygenase system

Cited by 12 publications

References 18 publications

Enantioselective Epoxidation by Flavoprotein Monooxygenases Supported by Organic Solvents

Enantioselective Epoxidation by Flavoprotein Monooxygenases Supported by Organic Solvents

Biodiversity of Microorganisms Colonizing the Surface of Polystyrene Samples Exposed to Different Aqueous Environments

De Novo Design, Solution Characterization, and Crystallographic Structure of an Abiological Mn–Porphyrin-Binding Protein Capable of Stabilizing a Mn(V) Species

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