Direct electrochemical generation of catalytically competent oxyferryl species of classes i and p dye decolorizing peroxidases

This work introduces a novel way to obtain catalytically competent oxyferryl species for two different dye-decolorizing peroxidases (DyPs) in the absence of H2O2 or any other peroxide by simply applying a reductive electrochemical potential under aerobic conditions. UV-vis and resonance Raman spectroscopies show that this method yields long-lived compounds II and I for the DyPs from Bacillus subtilis (BsDyP; Class I) and Pseudomonas putida (PpDyP; Class P), respectively. Both electrochemically generated high valent intermediates are able to oxidize ABTS at both acidic and alkaline pH. Interestingly, the electrocatalytic efficiencies obtained at pH 7.6 are very similar to the values recorded for regular catalytic ABTS/H2O2 assays at the optimal pH of the enzymes, ca. 3.7. These findings pave the way for the design of DyP-based electrocatalytic reactors operable in an extended pH range without the need of harmful reagents such as H2O2.Fil: Scocozza, Magali Franca. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química, Física de los Materiales, Medioambiente y Energía. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química, Física de los Materiales, Medioambiente y Energía; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Inorgánica, Analítica y Química Física; ArgentinaFil: Martins, Lígia O.. Universidade Nova de Lisboa; PortugalFil: Murgida, Daniel Horacio. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química, Física de los Materiales, Medioambiente y Energía. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química, Física de los Materiales, Medioambiente y Energía; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Inorgánica, Analítica y Química Física; Argentin

A wide array of lignin-related phenolics are oxidized by an evolved bacterial dye-decolourising peroxidase

Lignin is the second most abundant natural polymer next to cellulose and by far the largest renewable source of aromatic compounds on the planet. Dye-decolourising peroxidases (DyPs) are biocatalysts with immense potential in lignocellulose biorefineries to valorize emerging lignin building blocks for environmentally friendly chemicals and materials. This work investigates the catalytic potential of the engineered PpDyP variant 6E10 for the oxidation of 24 syringyl, guaiacyl and hydroxybenzene lignin-phenolic derivatives. Variant 6E10 exhibited up to 100-fold higher oxidation rates at pH 8 for all the tested phenolic substrates compared to the wild-type enzyme and other acidic DyPs described in the literature. The main products of reactions were dimeric isomers with molecular weights of (2 × MWsubstrate - 2 H). Their structure depends on the substitution pattern of the aromatic ring of substrates, i.e., of the coupling possibilities of the primarily formed radicals upon enzymatic oxidation. Among the dimers identified were syringaresinol, divanillin and diapocynin, important sources of structural scaffolds exploitable in medicinal chemistry, food additives and polymers.info:eu-repo/semantics/publishedVersio

An enzymatic route to a benzocarbazole framework using bacterial CotA laccase

The CotA laccase-catalysed oxidation of the meta, para-disubstituted arylamine 2,4-diaminophenyldiamine delivers, under mild reaction conditions, a benzocarbazole derivative (1) (74% yield), a key structural motif of a diverse range of applications. This work extends the scope of aromatic frameworks obtained using these enzymes and represents a new efficient and clean method to construct in one step C-C and C-N bonds

Mechanisms underlying dioxygen reduction in laccases. Structural and modelling studies focusing on proton transfer

<p>Abstract</p> <p>Background</p> <p>Laccases are enzymes that couple the oxidation of substrates with the reduction of dioxygen to water. They are the simplest members of the multi-copper oxidases and contain at least two types of copper centres; a mononuclear T1 and a trinuclear that includes two T3 and one T2 copper ions. Substrate oxidation takes place at the mononuclear centre whereas reduction of oxygen to water occurs at the trinuclear centre.</p> <p>Results</p> <p>In this study, the CotA laccase from <it>Bacillus subtilis </it>was used as a model to understand the mechanisms taking place at the molecular level, with a focus in the trinuclear centre. The structures of the holo-protein and of the oxidised form of the apo-protein, which has previously been reconstituted <it>in vitro </it>with Cu(I), have been determined. The former has a dioxygen moiety between the T3 coppers, while the latter has a monoatomic oxygen, here interpreted as a hydroxyl ion. The UV/visible spectra of these two forms have been analysed in the crystals and compared with the data obtained in solution. Theoretical calculations on these and other structures of CotA were used to identify groups that may be responsible for channelling the protons that are needed for reduction of dioxygen to water.</p> <p>Conclusions</p> <p>These results present evidence that Glu 498 is the only proton-active group in the vicinity of the trinuclear centre. This strongly suggests that this residue may be responsible for channelling the protons needed for the reduction. These results are compared with other data available for these enzymes, highlighting similarities and differences within laccases and multicopper oxidases.</p

Acute Effects of Different Set Configurations on Neuromuscular, Metabolic, and Perceptual Responses in Young Women

International Journal of Exercise Science 16(4): 974-986, 2023. We compared neuromuscular, metabolic, and perceptual responses between different resistance training configurations in young women. In a counterbalanced randomized order, 13 young women performed the following protocols in separate sessions (sets x repetitions): traditional (TRAD): 5x10, 90-s of rest interval between sets; more frequent and shorter total rest (FSR): 10x5, 30-s of rest interval between sets. The sessions were composed of leg press exercise with the same intensity. Force (maximum voluntary isometric contraction [MVIC]) and metabolic (lactate concentration) responses were measured pre- and post-resistance training sessions. The rating of perceived exertion (RPE) was measured after each set. The internal training load was calculated using the session-RPE method. There was a significant reduction in the MVIC only after TRAD configuration (Effect size [ES] = 0.36). The lactate concentration increased in both conditions but was higher after TRAD (ES = 2.81) than FSR (ES = 1.23). The RPE has progressively increased in both configurations. On the other hand, the internal training load was lower in the FSR configuration. From our findings, we suggest that more frequent and shorter total rest is an effective strategy for maintaining the ability to produce force, generating less metabolic stress and lower perceived internal load in young women

Towards the rational biosynthesis of substituted phenazines and phenoxazinones by laccases

Laccases are multi-copper oxidases that oxidise a wide range of substrates including phenol and aniline derivatives, which could be further involved in coupling reactions leading to the formation of dimeric and trimeric structures. This paper describes the enzyme-mediated dimerisation of several ortho and meta, para-disubstituted aromatic amines into phenazine ("head-to-tail" dimers) and phenoxazinone chromophores. The redox properties of substituted aromatic amines were studied by cyclic voltammetry and the kinetic constants of CotA and Trametes versicolor laccases were measured for selected aromatic amines. The structure of novel enzymatically synthesised phenazine and phenoxazinone dyes using CotA laccase was assessed by NMR and MS. Overall our data show that this enzymatic green process is an efficient alternative to the classic chemical oxidation of aromatic amines and phenols, with an impact on the broad field of applications of these heterocyclic compounds

Methyl syringate: An efficient phenolic mediator for bacterial and fungal laccases

The aim of the present work is to provide insight into the mechanism of laccase reactions using syringyl-type mediators. We studied the pH dependence and the kinetics of oxidation of syringyl-type phenolics using the low CotA and the high redox potential TvL laccases. Additionally, the efficiency of these compounds as redox mediators for the oxidation of non-phenolic lignin units was tested at different pH values and increasing mediator/non-phenolic ratios. Finally, the intermediates and products of reactions were identified by LC-MS and H-1 NMR. These approaches allow concluding on the (1) mechanism involved in the oxidation of phenolics by bacterial laccases, (2) importance of the chemical nature and properties of phenolic mediators, (3) apparent independence of the enzyme's properties on the yields of non-phenolics conversion, (4) competitive routes involved in the catalytic cycle of the laccase-mediator system with several new C-O coupling type structures being proposed

A One-Pot, Whole-Cell Biocatalysis Approach for Vanillin Production using Lignin Oil

Vanillin is a popular and versatile flavor compound, almost entirely produced from petroleum-derived phenol by a multi-step chemical synthesis. The process is hazardous to the environment and unsustainable for its fossil oil usage. Therefore, developing environmentally friendly, efficient, and sustainable routes to biobased vanillin is essential. Here, we report on vanillin production from 4-n-propylguaiacol (4PG), one of the main components in lignin oil obtained through reductive catalytic fractionation (RCF) of soft wood, by employing recombinant Escherichia coli cells. Conversion is based on the expression of two engineered oxidative enzymes: a 4-n-propylguaiacol oxidase and an isoeugenol dioxygenase. A high yield of vanillin, 66% from 4PG in RCF lignin oil was achieved through rounds of optimisation of the whole-cell conversion process. This high-performance strategy was readily scaled up to produce vanillin at an unprecedented 18% and 3% yield based on lignin oil and spruce wood respectively. The whole-cell bioconversion process shows good tolerance even at high loadings of starting material, showcasing the robustness and applicability of the employed biocatalysts. This work paves the way for further development towards the efficient production of high-titer biobased vanillin using depolymerised lignin as the feedstock.</p