More than Just a Fortuitous Sulfur Substitute in Redox Biology

FCT/MCTES is also acknowledged for the CEEC-Individual Program Contract (LBM). This work was also funded by DST–SERB for the CRG grant (file no CRG/2022/005673) and the Cluster University of Jammu for providing infrastructure facilities. Publisher Copyright: © 2023 by the authors.Living organisms use selenium mainly in the form of selenocysteine in the active site of oxidoreductases. Here, selenium’s unique chemistry is believed to modulate the reaction mechanism and enhance the catalytic efficiency of specific enzymes in ways not achievable with a sulfur-containing cysteine. However, despite the fact that selenium/sulfur have different physicochemical properties, several selenoproteins have fully functional cysteine-containing homologues and some organisms do not use selenocysteine at all. In this review, selected selenocysteine-containing proteins will be discussed to showcase both situations: (i) selenium as an obligatory element for the protein’s physiological function, and (ii) selenium presenting no clear advantage over sulfur (functional proteins with either selenium or sulfur). Selenium’s physiological roles in antioxidant defence (to maintain cellular redox status/hinder oxidative stress), hormone metabolism, DNA synthesis, and repair (maintain genetic stability) will be also highlighted, as well as selenium’s role in human health. Formate dehydrogenases, hydrogenases, glutathione peroxidases, thioredoxin reductases, and iodothyronine deiodinases will be herein featured.publishersversionpublishe

Enzyme-inspired dry-powder polymeric catalyst for green and fast pharmaceutical manufacturing processes

Funding Information: The authors thank financial support from Fundação para a Ciência e a Tecnologia , Ministério da Ciência, Tecnologia e Ensino Superior (FCT/MCTES Portugal), through project PTDC/EQU-EQU/32473/2017 , a Principal Investigator contract IF/00915/2014 (T.C.), and a doctoral grant SFRH/BDE/51907/2012 , a partnership from FCT/MCTES and the pharmaceutical company HOVIONE (R.V.). L.B.M. would like to acknowledge for FCT/MCTES funding with reference CEECIND/03810/2017. The NMR spectrometers in LabNMR@Cenimat are part of the National NMR Facility, supported by FCT (ROTEIRO/0031/2013 - PINFRA/22161/2016 ), co-financed by FEDER through COMPETE 2020, POCI, and PORL and FCT through PIDDAC ( POCI-01-0145-FEDER-007688 ; UID/CTM/50025/2020-2023 ). The Associate Laboratory Research Unit for Green Chemistry - Clean Technologies and Processes - LAQV is financed by national funds from FCT/MCTES ( UIDB/QUI/50006/2020 ) and cofunded by the ERDF under the PT2020 Partnership Agreement ( POCI-01-0145-FEDER-007265 ). We also acknowledge Dr. Luz Fernandes, REQUIMTE analytical services, for GC analysis. Publisher Copyright: © 2022Catalysis in pharma manufacturing processes is typically homogeneous, expensive and with hard catalyst recovery/regeneration. Herein an enzyme-inspired dry-powder molecularly imprinted polymeric (MIP) system was designed for fast, selective oxidation of a cholesterol derivative and easy catalyst regeneration. The strategy involved the synthesis of a template-monomer (T:M) complex followed by the crosslinked polymerization in supercritical carbon dioxide (scCO2). A 2,2,6,6-tetramethyl-1-piperidinyloxy (TEMPO)-MIP catalyst is obtained after the template cleavage from the matrix, and the oxidation of the N[sbnd]H groups turns available TEMPO moieties within the MIP. The oxidation of benzyl alcohol, 5α-cholestan-3β-ol and cholic acid was fast, in high yield and with selective oxidation capacity.publishersversionpublishe

Third-generation electrochemical biosensor based on nitric oxide reductase immobilized in a multiwalled carbon nanotubes/1-n-butyl-3-methylimidazolium tetrafluoroborate nanocomposite for nitric oxide detection

Nitric oxide (NO) has a crucial role in signaling and cellular physiology in humans. Herein, a novel third-generation biosensor based on the Marinobacter hydrocarbonoclasticus metalloenzyme (nitric oxide reductase (NOR)), responsible for the NO reduction in the denitrifying processes, was developed through the direct adsorption of a new nanocomposite (multiwalled carbon nanotubes (MWCNTs)/1-n-butyl-3-methylimidazolium tetrafluoroborate (BMIMBF4)/NOR) onto a pyrolytic graphite electrode (PGE) surface. The NOR direct electron transfer behavior (formal potential of -0.255 ± 0.003 V vs. Ag/AgCl) and electrocatalysis towards NO reduction (−0.68 ± 0.03 V vs. Ag/AgCl) of the PGE/[MWCNTs/BMIMBF4/NOR] biosensor were investigated in phosphate buffer at pH 6.0. Large enzyme loading (2.04 × 10−10 mol/cm2), acceptable electron transfer rate between NOR and the PGE surface (ks = 0.35 s-1), and high affinity for NO (Km = 2.17 μmol L-1) were observed with this biosensor composition. A linear response to NO concentration (0.23–4.76 μmol L-1) was perceived with high sensitivity (0.429 μA/μmolL-1), a detection limit of 0.07 μmol L-1, appropriate repeatability (9.1% relative standard deviations (RSD)), reproducibility (6.0–11% RSD) and 80–102% recoveries. The biosensor was stable during 1 month retaining 79–116% of its initial response. These data confirmed that NOR incorporated in the MWCNTs/BMIMBF4 nanocomposite can efficiently maintain its bioactivity paving a new and effective way for NO biosensing.FG thanks FCT/MCTES for the fellowship grant SFRH/BD/52502/2014, which is financed by national funds and co-financed by FSE. LBM thanks to FCT/MCTES for the CEEC-Individual 2017 Program Contract. This work was supported by the PTDC/BB-BQB/0129/2014 project (FCT/MCTES) and also by the Associate Laboratory Research Unit for Green Chemistry - Technologies and Processes Clean – LAQV, financed by national funds from FCT/MEC (UID/QUI/50006/2019) and co-financed by the ERDF under the PT2020 Partnership Agreement (POCI-01-0145-FEDER-007265). Funding through REQUIMTE project entitled “NOR-based biosensor for nitric oxide detection in biological and environmental samples” is also acknowledged. The financial support from the European Union (FEDER funds through COMPETE) and National Funds (Fundação para a Ciência e Tecnologia-FCT) through project PTDC/ASP-PES/29547/2017, by FCT/MEC with national funds and co-funded by FEDER, is also acknowledged.info:eu-repo/semantics/publishedVersio

Direct electrochemical reduction of carbon dioxide by a molybdenum-containing formate dehydrogenase

This work was supported by the Associate Laboratory for Green Chemistry -LAQV, with national funds from FCT/MCTES ( UID/QUI/50006/2019 ). LBM thanks to FCT/MCTES for the CEEC-Individual 2017 Program Contract.Formate dehydrogenase enzymes catalyse the reversible two-electron oxidation of formate to carbon dioxide. The class of metal-dependent formate dehydrogenases comprises prokaryotic enzymes holding redox-active centres and a catalytic site, containing either molybdenum or tungsten ion, that mediates the formate/carbon dioxide interconversion. The carbon dioxide reduction is of a particular interest, since it may be a route for its atmospheric mitigation with the simultaneous production of added-value products, as formate-derived compounds. Recently, the periplasmic formate dehydrogenase from Desulfovibrio desulfuricans, a molybdenum-containing enzyme, was proven to be an efficient enzyme for the CO 2 reduction to formate. In this work, the immobilized formate dehydrogenase isolated from Desulfovibrio desulfuricans direct electrochemical behaviour was attained in the presence and absence of substrates and the formal potentials associated with the catalytic centre transitions were determined in non-turnover conditions. The enzyme catalytic activity towards carbon dioxide reduction was observed using direct electrochemical methods.authorsversionpublishe

Química de coordenação e Biologia: controlo da actividade enzimática por alteração da coordenação de centros metálicos catalíticos

Os autores agradecem o apoio financeiro concedido pela Fundação para a Ciência e Tecnologia/MEC (PTDC/QUI-BIQ/100366/2008, PTDC/BIA-PRO/109796/2009, PTDC/QUI-BIQ/116481/2010, e PEst-C/EQB/ LA0006/2013).publishersversionpublishe

Assessment of habitat suitability for common cockles in the Ria the Aveiro Lagoon under average and projected environmental conditions

The common cockle Cerastoderma edule is a widespread bivalve species inhabiting estuarine systems across the North East Atlantic, where it provides several ecosystem services, and represents a valuable fishery resource for local economies. However, anthropogenic pressure and more frequent extreme weather events threaten the resilience of the species. Spatially explicit information on species distribution is critical for the implementation of management and conservation practices. This study assessed the potential distribution of C. edule in the Ria de Aveiro by estimating the habitat suitability using an ensemble approach based on ecological niche modeling and recently developed hydrodynamic and water quality models to forecast both average and projected estuarine conditions. The models were developed for the summer of 2013 and spring of 2019 and potential range shifts in the species distribution were forecasted under projected environmental conditions: high and low estimates of freshwater discharge, a 2 °C increase in water temperature, and the combined effect of low freshwater discharge and increased water temperature. The results suggest that salinity, time of submersion, and current velocity play an important role in the distribution of cockles, and large areas were consistently classified with high habitat suitability. Increased freshwater discharge (both seasons) and low discharge coupled to increased temperature (spring) resulted in large decreases in suitable habitat. Conversely, low freshwater discharges and average (unchanged) temperatures increased the suitable habitat in the outermost regions of the Ria. The spatially explicit information provided contributes to a better understanding of the vulnerability of C. edule in the Ria de Aveiro to extreme weather events (e.g., droughts, river floods) and may support adaptive management strategies of the cockle fishery during these conditions. Moreover, this approach can be transferred to other estuarine ecosystems for which data describing the environmental conditions (e.g., derived from numerical models), and information about species presence are available (including data-poor species).publishe

Electroanalytical characterization of the direct Marinobacter hydrocarbonoclasticus nitric oxide reductase-catalysed nitric oxide and dioxygen reduction

Understanding the direct electron transfer processes between redox proteins and electrode surface is fundamental to understand the proteins mechanistic properties and for development of novel biosensors. In this study, nitric oxide reductase (NOR) extracted from Marinobacter hydrocarbonoclasticus bacteria was adsorbed onto a pyrolytic graphite electrode (PGE) to develop an unmediated enzymatic biosensor (PGE/NOR)) for characterization of NOR direct electrochemical behaviour and NOR electroanalytical features towards NO and O2. Square-wave voltammetry showed the reduction potential of all the four NOR redox centers: 0.095 ± 0.002, -0.108 ± 0.008, -0.328 ± 0.001 and -0.635 ± 0.004 V vs. SCE for heme c, heme b, heme b3 and non-heme FeB, respectively. The determined sensitivity (-4.00 × 10-8 ± 1.84 × 10-9 A/μM and - 2.71 × 10-8 ± 1.44 × 10-9 A/μM for NO and O2, respectively), limit of detection (0.5 μM for NO and 1.0 μM for O2) and the Michaelis Menten constant (2.1 and 7.0 μM for NO and O2, respectively) corroborated the higher affinity of NOR for its natural substrate (NO). No significant interference on sensitivity towards NO was perceived in the presence of O2, while the O2 reduction was markedly and negatively impacted (3.6 times lower sensitivity) by the presence of NO. These results clearly demonstrate the high potential of NOR for the design of innovative NO biosensors.FG and LBM thank FCT/MCTES for the fellowship grants SFRH/BD/52502/2014 and SFRH/BPD/111404/2015, respectively, which are financed by national funds and co-financed by FSE. CMC acknowledges FCT-MCTES funding through project PTDC/BBB-BQB/0129/2014 (FCT/MCTES). This work was supported by the REQUIMTE, which is financed by national funds from FCT/MCTES (UID/QUI/50006/2013 and UID/Multi/04378/2013) and co-financed by the ERDF under the PT2020 Partnership Agreement (POCI-01-0145-FEDER-007265 and POCI-01-0145-FEDER-007728), and also by the PTDC/BB-BQB/0129/2014 project (FCT/MCTES). Funding through REQUIMTE project entitled “NOR-based biosensor for nitric oxide detection in biological and environmental samples” is also acknowledged.info:eu-repo/semantics/publishedVersio

Nitric Oxide Detection Using Electrochemical Third-generation Biosensors - Based on Heme Proteins and Porphyrins

Nitric oxide radical (NO) is a signalling molecule involved in virtually all forms of life. Its relevance has been leading to the development of different analytical methodologies to assess the temporal and spatial fluxes of NO under the complex biological milieu. Third‐generation electrochemical biosensors are promising tools for in loco and in vivo NO quantification and, over the past years, heme proteins and porphyrins have been used in their design. Since there are some limitations with the biorecognition element directly adsorbed onto the electrode surface, nanomaterials (carbon nanotubes, gold nanoparticles, etc.) and polymers (cellulose, chitosan, nafion®, polyacrylamide, among others) have been explored to achieve high kinetics and better biosensor performance. In this review, a broad overview of the field of electrochemical third‐generation biosensors for NO electroanalysis is presented, discussing their main characteristics and aiming new outlooks and advances in this field.FG and LBM thank FCT/MCTES for the fellowship grants SFRH/BD/52502/2014 and SFRH/BPD/111404/ 2015, respectively, which are financed by national funds and co-financed by FSE. CMC acknowledges FCTMCTES funding through project PTDC/BBB-BQB/ 0129/2014 (FCT/MCTES). This work was supported by the PTDC/BB-BQB/0129/2014 project (FCT/MCTES) and also by the Associate Laboratory Research Unit for Green Chemistry – Technologies and Processes Clean – LAQV, financed by national funds from FCT/MEC (UID/QUI/50006/2013) and co-financed by the ERDF under the PT2020 Partnership Agreement (POCI-01- 0145-FEDER-007265). Funding through REQUIMTE project entitled “NOR-based biosensor for nitric oxide detection in biological and environmental samples” is also acknowledged.info:eu-repo/semantics/publishedVersio

Biosensor for direct bioelectrocatalysis detection of nitric oxide using nitric oxide reductase incorporated in carboxylated single-walled carbon nanotubes/lipidic 3 bilayer nanocomposite

An enzymatic biosensor based on nitric oxide reductase (NOR; purified from Marinobacter hydrocarbonoclasticus) was developed for nitric oxide (NO) detection. The biosensor was prepared by deposition onto a pyrolytic graphite electrode (PGE) of a nanocomposite constituted by carboxylated single-walled carbon nanotubes (SWCNTs), a lipidic bilayer [1,2-di-(9Z-octadecenoyl)-sn-glycero-3-phosphoethanolamine (DOPE), 1,2-di-(9Z-octadecenoyl)-3-trimethylammonium-propane (DOTAP), 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-polyethylene glycol (DSPE-PEG)] and NOR. NOR direct electron transfer and NO bioelectrocatalysis were characterized by several electrochemical techniques. The biosensor development was also followed by scanning electron microscopy and Fourier transform infrared spectroscopy. Improved enzyme stability and electron transfer (1.96 × 10-4 cm.s-1 apparent rate constant) was obtained with the optimum SWCNTs/(DOPE:DOTAP:DSPE-PEG)/NOR) ratio of 4/2.5/4 (v/v/v), which biomimicked the NOR environment. The PGE/[SWCNTs/(DOPE:DOTAP:DSPE-PEG)/NOR] biosensor exhibited a low Michaelis-Menten constant (4.3 μM), wide linear range (0.44-9.09 μM), low detection limit (0.13 μM), high repeatability (4.1% RSD), reproducibility (7.0% RSD), and stability (ca. 5 weeks). Selectivity tests towards L-arginine, ascorbic acid, sodium nitrate, sodium nitrite and glucose showed that these compounds did not significantly interfere in NO biosensing (91.0 ± 9.3%-98.4 ± 5.3% recoveries). The proposed biosensor, by incorporating the benefits of biomimetic features of the phospholipid bilayer with SWCNT's inherent properties and NOR bioelectrocatalytic activity and selectivity, is a promising tool for NO.FG thanks to Fundacão para a Ciência e a Tecnologia, MCTES (FCT/MCTES) for the fellowship grant SFRH/BD/52502/2014, which is financed by national funds and co-financed by FSE. LBM thanks to FCT/MCTES for the CEEC-Individual 2017 Program Contract. This work was supported by the PTDC/BB-BQB/0129/2014 project (FCT/MCTES), by FCT/MEC with national funds and co-funded by FEDER, and also by the Associate Laboratory Research Unit for Green Chemistry - Technologies and Processes Clean – LAQV, financed by national funds from FCT/MCTES (UID/QUI/50006/2019). The financial support from the European Union (FEDER funds through COMPETE) and National Funds (Fundação para a Ciência e Tecnologia-FCT) through project PTDC/ASP-PES/29547/2017, by FCT/MEC with national funds and co-funded by FEDER, is also acknowledged. J.A. Loureiro post-doc grant was supported by NORTE-01-0145-FEDER-000005 – LEPABE-2 ECO-INNOVATION, from North Portugal Regional Operational Program (NORTE 2020), under the Portugal 2020 Partnership Agreement, through the European Regional Development Fund (ERDF).info:eu-repo/semantics/publishedVersio