ORCID Profile
0000-0001-5264-464X
Current Organisations
University of Leicester
,
University of Oxford
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Publisher: Royal Society of Chemistry (RSC)
Date: 2019
DOI: 10.1039/C9DT00924H
Abstract: Iron–sulfur clusters are exceptionally tuneable protein cofactors, and as one of their many roles they are involved in biological responses to nitrosative stress.
Publisher: Royal Society of Chemistry (RSC)
Date: 2016
DOI: 10.1039/C6CC05955D
Abstract: Adsorbed formate is observed on a supported Pt nanoparticle for the first time during formic acid electro-oxidation.
Publisher: American Chemical Society (ACS)
Date: 02-08-2018
DOI: 10.1021/JACS.8B04798
Abstract: Catalytic long-range proton transfer in [NiFe]-hydrogenases has long been associated with a highly conserved glutamate (E) situated within 4 Å of the active site. Substituting for glutamine (Q) in the O
Publisher: Royal Society of Chemistry (RSC)
Date: 2021
DOI: 10.1039/D1SC01734A
Abstract: Electrochemically-coupled IR microspectroscopy of single crystals provides insight into proton-coupled electron transfer in [NiFe] hydrogenase.
Publisher: American Chemical Society (ACS)
Date: 07-03-2017
Publisher: Royal Society of Chemistry (RSC)
Date: 2014
DOI: 10.1039/C4FD00058G
Abstract: We present a study of electrocatalysis by an enzyme adsorbed on a range of carbon materials, with different size, surface area, morphology and graphitic structure, which are either commercially available or prepared via simple, established protocols. We choose as our model enzyme the hydrogenase I from E. coli (Hyd-1), which is an active catalyst for H 2 oxidation, is relatively robust and has been demonstrated in H 2 fuel cells and H 2 -driven chemical synthesis. The carbon materials were characterised according to their surface area, surface morphology and graphitic character, and we use the electrocatalytic H 2 oxidation current for Hyd-1 adsorbed on these materials to evaluate their effectiveness as enzyme electrodes. Here, we show that a variety of carbon materials are suitable for adsorbing hydrogenases in an electroactive configuration. This unified study provides insight into selection and design of carbon materials for study of redox enzymes and different applications of enzyme electrocatalysis.
Publisher: American Chemical Society (ACS)
Date: 05-08-2014
DOI: 10.1021/JA505291J
Abstract: We use infrared spectroscopy to demonstrate the critical role that trace O2 plays in determining the products formed when a [2Fe2S] cluster protein reacts with nitric oxide (NO). The observed importance of O2 may have physiological relevance, as many pathogens sense NO using iron-sulfur proteins and will be exposed to NO in an aerobic environment during a mammalian immune response. We show that the [2Fe2S]-containing spinach ferredoxin I undergoes reaction with NO at pH 6.0, with the proportion of protein-bound Roussin's Red Ester compared to the dinitrosyl iron complex product favored by trace O2. Roussin's Red Ester is also favored on nitrosylation in the presence of the thiolate scavenging reagent, iodoacetamide, suggesting that the role of O2 is in oxidative sequestration of cysteine thiolates. Infrared spectroscopy has been overlooked as a tool for studying iron-sulfur protein nitrosylation despite the fact that there exists a wealth of infrared spectroscopic data on small-molecule nitrosyl clusters which serve as models for the identification of protein-bound nitrosyl clusters.
Publisher: American Chemical Society (ACS)
Date: 10-07-2015
Abstract: The regulatory hydrogenase (RH) from Ralstonia eutropha acts as the H2-sensing unit of a two-component system that regulates biosynthesis of the energy conserving hydrogenases of the organism according to the availability of H2. The H2 oxidation activity, which was so far determined in vitro with artificial electron acceptors, has been considered to be insensitive to O2 and CO. It is assumed that bulky isoleucine and phenylalanine amino acid residues close to the NiFe active site "gate" gas access, preventing molecules larger than H2 interacting with the active site. We have carried out sensitive electrochemical measurements to demonstrate that O2 is in fact an inhibitor of H2 oxidation by the RH, and that both H(+) reduction and H2 oxidation are inhibited by CO. Furthermore, we have demonstrated that the inhibitory effect of O2 arises due to interaction of O2 with the active site. Using protein film infrared electrochemistry (PFIRE) under H2 oxidation conditions, in conjunction with solution infrared measurements, we have identified previously unreported oxidized inactive and catalytically active reduced states of the RH active site. These findings suggest that the RH has a rich active site chemistry similar to that of other NiFe hydrogenases.
Publisher: Royal Society of Chemistry (RSC)
Date: 2016
DOI: 10.1039/C6FD90069K
Publisher: Royal Society of Chemistry (RSC)
Date: 2021
DOI: 10.1039/D1DT02219A
Abstract: The distribution of redox species in single crystals of [FeFe]-hydrogenase can be tuned electrochemically. All catalytic intermediates are observed by simultaneous infrared microspectroscopy, providing a roadmap for enriching specific redox states.
Publisher: Wiley
Date: 29-04-2015
Publisher: Royal Society of Chemistry (RSC)
Date: 2023
DOI: 10.1039/D2FD00170E
Abstract: We associate potentials with the binding of substrates and inhibitors, H + , C 2 H 2 , CO and MeNC, to nitrogenase MoFe protein by coupling electrochemical control with gas chromatography analysis and in situ infrared spectroscopy.
Publisher: Royal Society of Chemistry (RSC)
Date: 2017
DOI: 10.1039/C6SC02860H
Abstract: Electrochemical control over nitrogenase allows us to examine electrocatalytic proton reduction and potential-triggered CO inhibition using infrared spectroscopy.
Publisher: Royal Society of Chemistry (RSC)
Date: 2017
DOI: 10.1039/C7CC02591B
Abstract: We manipulate and verify the redox state of single metalloprotein crystals by combining electrochemical control with synchrotron infrared microspectroscopy.
Publisher: Royal Society of Chemistry (RSC)
Date: 2023
DOI: 10.1039/D2SC05641K
Abstract: Changing the conserved active-site arginine of [NiFe]-hydrogenases into a lysine greatly lowers the rates of catalytic H 2 activation in each direction and results in the extremely tight binding of a diatomic ligand.
Publisher: American Chemical Society (ACS)
Date: 07-08-2017
Publisher: Royal Society of Chemistry (RSC)
Date: 2013
DOI: 10.1039/C3CP00119A
Abstract: We report a versatile infrared spectroscopic method for studying redox chemistry of metalloproteins, and demonstrate for the first time electrochemically-induced changes to the active site of the regulatory [NiFe]-hydrogenase from Ralstonia eutropha. A carbon particle network working electrode allows control over a wide potential window without the need for solution mediators.
Publisher: American Chemical Society (ACS)
Date: 15-06-2016
Publisher: Royal Society of Chemistry (RSC)
Date: 2012
DOI: 10.1039/C1CC14826E
Abstract: Pyrolytic graphite particles modified with hydrogenase and an NAD(+)/NADH cycling enzyme provide a modular heterogeneous catalyst system for regeneration of oxidised or reduced nicotinamide cofactors using H(2) and H(+) as electron source or sink. Particles can be tuned for cofactor supply under different conditions by appropriate choice of hydrogenase.
Publisher: American Chemical Society (ACS)
Date: 11-2019
DOI: 10.1021/ACS.ACCOUNTS.9B00293
Abstract: Achieving a unified understanding of the mechanism of a multicenter redox enzyme such as [NiFe] hydrogenase is complicated by difficulties in reconciling information obtained by using different techniques and on s les in different physical forms. Measurements of the activity of the enzyme, and of factors which perturb activity, are generally carried out using biochemical assays in solution or with electrode-immobilized enzymes using protein film electrochemistry (PFE). Conversely, spectroscopy aimed at reporting on features of the metalloclusters in the enzyme, such as electron paramagnetic resonance (EPR) or X-ray absorption spectroscopy (XAS), is often conducted on frozen s les and is thus difficult to relate to catalytically relevant states as information about turnover and activity has been lost. To complicate matters further, most of our knowledge of the atomic-level structure of metalloenzymes comes from X-ray diffraction studies in the solid, crystalline state, which are again difficult to link to turnover conditions. Taking [NiFe] hydrogenases as our case study, we show here how it is possible to apply infrared (IR) spectroscopic s ling approaches to unite direct spectroscopic study with catalytic turnover. Using a method we have named protein film IR electrochemistry (PFIRE), we reveal the steady-state distribution of intermediates during catalysis and identify catalytic "bottlenecks" introduced by site-directed mutagenesis. We also show that it is possible to study dynamic transitions between active site states of enzymes in single crystals, uniting solid state and solution spectroscopic information. In all of these cases, the spectroscopic data complement and enhance interpretation of purely activity-based measurements by providing direct chemical insight that is otherwise hidden. The [NiFe] hydrogenases possess a bimetallic [NiFe] active site, coordinated by CO and CN
Publisher: Royal Society of Chemistry (RSC)
Date: 2012
DOI: 10.1039/C1CC15871F
Abstract: This article reviews recent developments in spectroscopic analysis of electrode-immobilised enzymes under direct, unmediated electrochemical control. These methods unite the suite of spectroscopic methods available for characterisation of structural, electronic and coordination changes in proteins with the exquisite control over complex redox enzymes that can be achieved in protein film electrochemistry in which immobilised protein molecules exchange electrons directly with an electrode. This combination is particularly powerful in studies of highly active enzymes where redox states can be controlled even under fast electrocatalytic turnover. We examine ex les in which UV-visible, IR, Raman and MCD spectroscopy have been combined with direct electrochemistry to probe redox-dependent chemistry, and consider future opportunities for 'direct' spectroelectrochemistry of immobilised enzymes.
Location: United Kingdom of Great Britain and Northern Ireland
Location: United Kingdom of Great Britain and Northern Ireland
No related grants have been discovered for Philip Ash.