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J W Peters

Publications and source records attributed to J W Peters.

At least 19 recordsLinked to original sources
PubMed2005

Biomimetic synthesis of a H2 catalyst using a protein cage architecture.

A biomimetic approach has been used to develop an artificial hydrogenase that catalyses the efficient reduction of protons producing hydrogen gas. Analogous to the unique biological metal clusters found in hydrogenase enzymes, the engineered active sites are small, well-defined Pt clusters deposited on the interior of a heat shock protein cage architecture with stoichiometries of 150 to 1000 Pt per protein cage. The proton reduction reaction is driven by visible light through a coupled reaction with Ru(bpy)3(2+) and methyl viologen as an electron-transfer mediator. Hydrogen production rates are comparable to those of hydrogenase on a per protein basis and exceed production rates of other reported Pt-based catalysts. These results demonstrate the utility of a biomimetic approach toward addressing the needs of hydrogen production.

Archaeal Proteins↗
PubMed2001

Mechanistic features and structure of the nitrogenase alpha-Gln195 MoFe protein.

EPR signals observed under CO and C(2)H(2) during nitrogenase turnover were investigated for the alpha-Gln(195) MoFe protein, an altered form for which the alpha-His(195) residue has been substituted by glutamine. Under CO, samples show S = 1/2 hi- and lo-CO EPR signals identical to those recognized for the wild-type protein, whereas the S = 3/2 signals generated under high CO/high flux conditions differ. Previous work has revealed that the EPR spectrum generated under C(2)H(2) exhibits a signal (S(EPR1)) originating from the FeMo-cofactor having two or more bound C(2)H(2) adducts and a second signal (S(EPR2)) arising from a radical species [Sørlie, M., Christiansen, J., Dean, D. R., and Hales, B. J. (1999) J. Am. Chem. Soc. 121, 9457-9458]. Pressure-dependent studies show that the intensity of these signals has a sigmoidal dependency at low pressures and maximized at 0.1 atm C(2)H(2) with a subsequent decrease in steady-state intensity at higher pressures. Analogous signals are not recognized for the wild-type MoFe protein. Analysis of the principal g-factors of S(EPR2) suggests that it either represents an unusual metal cluster or is a carboxylate centered radical possibly originating from homocitrate. Both S(EPR1) and S(EPR2) exhibit similar relaxation properties that are atypical for S = 1/2 signals originating from Fe-S clusters or radicals and indicate a coupled relaxation pathway. The alpha-Gln(195) MoFe protein also exhibits these signals when incubated under turnover conditions in the presence of C(2)H(4). Under these conditions, additional inflections in the g 4-6 region assigned to ground-state transitions of an S = 3/2 spin system are also recognized and assigned to turnover states of the MoFe protein without C(2)H(4) bound. The structure of alpha-Gln(195) was crystallographically determined and found to be virtually identical to that of the wild-type MoFe protein except for replacement of an NuH-S hydrogen bond interaction between FeMo-cofactor and the imidazole side chain of alpha-His(195) by an analogous interaction involving Gln.

Amino Acid Substitution↗
PubMed2001

MgATP-Bound and nucleotide-free structures of a nitrogenase protein complex between the Leu 127 Delta-Fe-protein and the MoFe-protein.

A mutant form of the nitrogenase iron protein with a deletion of residue Leu 127, located in the switch II region of the nucleotide binding site, forms a tight, inactive complex with the nitrogenase molybdenum iron (MoFe) protein in the absence of nucleotide. The structure of this complex generated with proteins from Azotobacter vinelandii (designated the L127Delta-Av2-Av1 complex) has been crystallographically determined in the absence of nucleotide at 2.2 A resolution and with bound MgATP (introduced by soaking) at 3.0 A resolution. As observed in the structure of the complex between the wild-type A. vinelandii nitrogenase proteins stabilized with ADP.AlF(4-), the most significant conformational changes in the L127Delta complex occur in the Fe-protein component. While the interactions at the interface between the MoFe-protein and Fe-proteins are conserved in the two complexes, significant differences are evident at the subunit-subunit interface of the dimeric Fe-proteins, with the L127Delta-Av2 structure having a more open conformation than the wild-type Av2 in the complex stabilized by ADP.AlF(4-). Addition of MgATP to the L127Delta-Av2-Av1 complex results in a further increase in the separation between Fe-protein subunits so that the structure more closely resembles that of the wild-type, nucleotide-free, uncomplexed Fe-protein, rather than the Fe-protein conformation in the ADP.AlF(4-) complex. The L127Delta mutation precludes key interactions between the Fe-protein and nucleotide, especially, but not exclusively, in the region corresponding to the switch II region of G-proteins, where the deletion constrains Gly 128 and Asp 129 from forming hydrogen bonds to the gamma-phosphate and activating water for attack on this group, respectively. These alterations account for the inability of this mutant to support mechanistically productive ATP hydrolysis. The ability of the L127Delta-Av2-Av1 complex to bind MgATP demonstrates that dissociation of the nitrogenase complex is not required for nucleotide binding.

Adenosine Triphosphate↗
PubMed2001

Crystallization and preliminary X-ray analysis of a NADPH 2-ketopropyl-coenzyme M oxidoreductase/carboxylase.

NADPH 2-ketopropyl-coenzyme M (2-mercaptoethanesulfonate) oxidoreductase/carboxylase is the terminal enzyme in a metabolic pathway that results in the conversion of propylene to the central metabolite acetoacetate. This enzyme is an FAD-containing enzyme that is a member of the NADPH:disulfide oxidoreductase family of enzymes and catalyzes the cleavage and carboxylation of 2-ketopropyl-coenzyme M to form acetoacetate and coenzyme M. Crystallization trials have revealed that the highest diffraction quality crystals (better that 2.0 A resolution) could be achieved when the substrate or product of the reaction was added to the enzyme in a stoichiometric excess.

Crystallization↗
PubMed2000

Insights into nucleotide signal transduction in nitrogenase: structure of an iron protein with MgADP bound.

Coupling the energy of nucleoside triphosphate binding and hydrolysis to conformational changes is a common mechanism for a number of proteins with disparate cellular functions, including those involved in DNA replication, protein synthesis, and cell differentiation. Unique to this class of proteins is the dimeric Fe protein component of nitrogenase in which the binding and hydrolysis of MgATP controls intermolecular electron transfer and reduction of nitrogen to ammonia. In the work presented here, the MgADP-bound (or "off") conformational state of the nitrogenase Fe protein has been captured and a 2.15 A resolution X-ray crystal structure is presented. The structure described herein reveals likely mechanisms for long-range communication from the nucleotide-binding sites for controlling the affinity of association with the MoFe protein component. Two pathways, termed switches I and II, appear to be integral to this nucleotide signal transduction mechanism. In addition, the structure provides the basis for the changes in the biophysical properties of the [4Fe-4S] cluster observed when Fe protein binds nucleotides. The structure of the MgADP-bound Fe protein provides important insights into the respective contributions of nucleotide interaction and complex formation in defining the conformational states that are the keys to nitrogenase catalysis.

Adenosine Diphosphate↗
PubMed2000

Reversible carbon monoxide binding and inhibition at the active site of the Fe-only hydrogenase.

Carbon monoxide binding and inhibition have been investigated by electron paramagnetic resonance (EPR) spectroscopy in solution and in crystals of structurally described states of the Fe-only hydrogenase (CpI) from Clostridium pasteurianum. Simulation of the EPR spectrum of the as-isolated state indicates that the main component of the EPR spectrum consists of the oxidized state of the "H cluster" and components due to reduced accessory FeS clusters. Addition of carbon monoxide to CpI in the presence of dithionite results in the inhibition of hydrogen evolution activity, and a characteristic axial EPR signal [g(eff(1)), g(eff(2)), and g(eff(3)) = 2.0725, 2.0061, and 2.0061, respectively] was observed. Hydrogen evolution activity was restored by successive sparging with hydrogen and argon and resulted in samples that exhibited the native oxidized EPR signature that could be converted to the reduced form upon addition of sodium dithionite and hydrogen. To examine the relationship between the spectroscopically defined states of CpI and those observed structurally by X-ray crystallography, we have examined the CpI crystals using EPR spectroscopy. EPR spectra of the crystals in the CO-bound state exhibit the previously described axial signal associated with CO binding. The results indicate that the addition of carbon monoxide to CpI results in a single reversible carbon monoxide-bound species characterized by loss of enzyme activity and the distinctive axial EPR signal.

Binding Sites↗
PubMed2000

Modulating the midpoint potential of the [4Fe-4S] cluster of the nitrogenase Fe protein.

Protein-bound [FeS] clusters function widely in biological electron-transfer reactions, where their midpoint potentials control both the kinetics and thermodynamics of these reactions. The polarity of the protein environment around [FeS] clusters appears to contribute largely to modulating their midpoint potentials, with local protein dipoles and water dipoles largely defining the polarity. The function of the [4Fe-4S] cluster containing Fe protein in nitrogenase catalysis is, at least in part, to serve as the nucleotide-dependent electron donor to the MoFe protein which contains the sites for substrate binding and reduction. The ability of the Fe protein to function in this manner is dependent on its ability to adopt the appropriate conformation for productive interaction with the MoFe protein and on its ability to change redox potentials to provide the driving force required for electron transfer. Phenylalanine at position 135 is located near the [4Fe-4S] cluster of nitrogenase Fe protein and has been suggested by amino acid substitution studies to participate in defining both the midpoint potential and the nucleotide-induced changes in the [4Fe-4S] cluster. In the present study, the crystal structure of the Azotobacter vinelandii nitrogenase Fe protein variant having phenylalanine at position 135 substituted by tryptophan has been determined by X-ray diffraction methods and refined to 2.4 A resolution. A comparison of available Fe protein structures not only provides a structural basis for the more positive midpoint potential observed in the tryptophan substituted variant but also suggests a possible general mechanism by which the midpoint potential could be controlled by nucleotide interactions and nitrogenase complex formation.

Adenosine Diphosphate↗
PubMed2000

A novel FeS cluster in Fe-only hydrogenases.

Many microorganisms can use molecular hydrogen as a source of electrons or generate it by reducing protons. These reactions are catalysed by metalloenzymes of two types: NiFe and Fe-only hydrogenases. Here, we review recent structural results concerning the latter, putting special emphasis on the characteristics of the active site.

Amino Acid Sequence↗
PubMed1999

Binding of exogenously added carbon monoxide at the active site of the iron-only hydrogenase (CpI) from Clostridium pasteurianum.

A site for the binding of exogenously added carbon monoxide has been identified at the active site of the Fe-only hydrogenase (CpI) from Clostridium pasteurianum. The binding and inhibition of carbon monoxide have been exploited in biochemical and spectroscopic studies to gain mechanistic insights. In the present study, we have taken advantage of the ability to generate an irreversibly carbon monoxide bound state of CpI. The crystallization and structural characterization of CpI inhibited in the presence of carbon monoxide indicates the addition of a single molecule of carbon monoxide. The ability to generate crystals of the carbon monoxide bound state of the hydrogenase that are isomorphous to those of the native enzyme has allowed for a direct comparison of the crystallographic data and an unambiguous identification of the site of carbon monoxide binding at the active site of CpI. Carbon monoxide binds to an Fe atom of the 2Fe subcluster at the site of a terminally bound water molecule in the as crystallized native state of CpI that has been previously suggested to be a potential site of reversible hydrogen oxidation. Binding of carbon monoxide at this site results in an active site that is coordinately saturated with strong ligands (S, CO, and CN), providing a rational potential mechanism for inhibition of reversible hydrogen oxidation at the active site of CpI.

Bacterial Proteins↗
PubMed1999

Structure and mechanism of iron-only hydrogenases.

The recent elucidation of the structures of iron-only hydrogenases from the microorganisms Clostridium pasteurianum and Desulfovibrio desulfuricans has revealed that the presumed site of reversible hydrogen oxidation exists as a unique, protein-associated organometallic prosthetic group. Details of the hydrogenase structures provide insight into the chemical mechanism of this highly evolved catalyst.

Binding Sites↗
PubMed1999

Patient controlled analgesia in children and adolescents: a randomized controlled trial.

In children, patient controlled analgesia (PCA) and continuous infusion (CI) of morphine are well established methods of relieving postoperative pain. This study was designed to assess the efficacy of PCA plus background infusion (BI) (15 microg x kg(-1) x h(-1) and bolus doses of 15 microg x kg(-1) with a lock-out interval of 10 min) with CI (20 to 40 microg x kg(-1) x h(-1)) in terms of analgesia, morphine needs and side-effects. A stratified randomized controlled trial was carried out. 47 children aged 5-18 years undergoing major elective lower/upper abdominal or spinal surgery were allocated. The magnitude of surgery was assessed by the Severity of Surgical Stress scoring (SSS) system. Pain was assessed by self-report every three h. Side-effects compatible with morphine as well as morphine consumption were recorded. Morphine consumption was significantly increased in the PCA group compared with the CI group. Moreover, morphine consumption was associated with SSS, independent of the technique of administration. There were no significant differences between groups in pain scores or in the incidence of side-effects.

Abdomen↗
PubMed1998

X-ray crystal structure of the Fe-only hydrogenase (CpI) from Clostridium pasteurianum to 1.8 angstrom resolution.

A three-dimensional structure for the monomeric iron-containing hydrogenase (CpI) from Clostridium pasteurianum was determined to 1.8 angstrom resolution by x-ray crystallography using multiwavelength anomalous dispersion (MAD) phasing. CpI, an enzyme that catalyzes the two-electron reduction of two protons to yield dihydrogen, was found to contain 20 gram atoms of iron per mole of protein, arranged into five distinct [Fe-S] clusters. The probable active-site cluster, previously termed the H-cluster, was found to be an unexpected arrangement of six iron atoms existing as a [4Fe-4S] cubane subcluster covalently bridged by a cysteinate thiol to a [2Fe] subcluster. The iron atoms of the [2Fe] subcluster both exist with an octahedral coordination geometry and are bridged to each other by three non-protein atoms, assigned as two sulfide atoms and one carbonyl or cyanide molecule. This structure provides insights into the mechanism of biological hydrogen activation and has broader implications for [Fe-S] cluster structure and function in biological systems.

Amino Acid Sequence↗
PubMed1998

Safety and efficacy of semi-closed circle ventilation in small infants.

The purpose of this clinical trial was to investigate the safety and efficacy of semi-closed circle ventilation of the Dräger anaesthesia ventilators (Cicero, Cato), using a fresh gas flow (FGF) of 600 ml.min-1. Twenty infants, weighing less than 6000 g, without cardiorespiratory abnormalities who required general anaesthesia of at least 30 min were included. The FGF was reduced to 600 ml.min-1 after 10 min of denitrogenation with a FGF of 4 to 61.min-1. The composition of the FGF (600 ml.min-1) was calculated as follows: oxygen necessary for consumption (60 ml.min-1) plus the remaining FGF in a 1:2 relationship for oxygen. The inspiratory nitrogen fraction was calculated to exclude accumulation. Inspiratory fractions of O2 and N2O plus inspiratory and endtidal CO2 partial pressures and noninvasive oxygen saturation were the control parameters. The gas concentrations (O2 and N2O) remained within safe limits. Hypoxic gas concentrations were not observed. Neither nitrogen nor CO2 accumulated in the circle system. In conclusion, low flow anaesthesia can be performed safely in infants under 6000 grams with the Dräger Cicero and Cato anaesthesia ventilators.

Anesthesia, Closed-Circuit↗
PubMed1997

[How to manage a rectal lesion in horses in clinical practice].

The most feared complication after a rectal examination of horses and ponies is to cause a tear of all the layers in the intraperitoneal part of the rectum. The description of the anatomy is an attempt to explain that the rectal tear is mostly located in the dorsal or dorsolateral part of the rectum. This article describes a relatively simple suture technique for the rectal tear by using the needle of Deschamp which is modified by Professor B. Huskamp. Furthermore there is a description of this method as treatment of two horses in practice circumstances. The issue whether causing a rectal tear should be regarded as a case of malpractice is discussed. Advises as prevention of rectal tears are given.

Animals↗
PubMed1997

Redox-dependent structural changes in the nitrogenase P-cluster.

The structure of the nitrogenase MoFe-protein from Azotobacter vinelandii has been refined to 2.0 A resolution in two oxidation states. EPR studies on the crystals indicate that the structures correspond to the spectroscopically assigned oxidized (P(OX)/M(OX)) and the native or dithionite-reduced (P(N)/M(N)) forms of the enzyme. Both MoFe-protein structures are essentially identical, with the exception of the P-cluster. The MoFe-protein P-cluster in each state is found to contain eight Fe and seven S atoms. Interconversion between the two redox states involves movement of two Fe atoms and an exchange of protein coordination for ligands supplied by a central S atom. In the oxidized P(OX) state, the cluster is coordinated by the protein through six cysteine ligands, Ser-beta188 O gamma, and the backbone amide of Cys-alpha88. In the native P(N) state, Ser-beta188 O gamma and the amide N of Cys-alpha88 no longer coordinate the cluster due to movement of their coordinated Fe atoms toward the central sulfur. Consequently, this central sulfur adopts a distorted octahedral environment with six surrounding Fe atoms. A previously described model of the P-cluster containing 8Fe-8S likely reflects the inappropriate modeling of a single structure to a mixture of these two P-cluster redox states. These observed redox-mediated structural changes of the P-cluster suggest a role for this cluster in coupling electron transfer and proton transfer in nitrogenase.

Aspergillus↗
PubMed1995

Involvement of the P cluster in intramolecular electron transfer within the nitrogenase MoFe protein.

Nitrogenase is the catalytic component of biological nitrogen fixation, and it is comprised of two component proteins called the Fe protein and MoFe protein. The Fe protein contains a single Fe4S4 cluster, and the MoFe protein contains two metallocluster types called the P cluster (Fe8S8) and FeMo-cofactor (Fe7S9Mo-homocitrate). During turnover, electrons are delivered one at a time from the Fe protein to the MoFe protein in a reaction coupled to component-protein association-dissociation and MgATP hydrolysis. Under conditions of optimum activity, the rate of component-protein dissociation is rate-limiting. The Fe protein's Fe4S4 cluster is the redox entity responsible for intermolecular electron delivery to the MoFe protein, and FeMo-cofactor provides the substrate reduction site. In contrast, the role of the P cluster in catalysis is not well understood although it is believed to be involved in accumulating electrons delivered from the Fe protein and brokering their intramolecular delivery to the substrate reduction site. A nitrogenase component-protein docking model, which is based on the crystallographic structures of the component proteins and which pairs the 2-fold symmetric surface of the Fe protein with the exposed surface of the MoFe protein's pseudosymmetric alpha beta interface, is now available. During component-protein interaction, this model places the P cluster between the Fe protein's Fe4S4 cluster and FeMo-cofactor, which implies that the P cluster is involved in mediating intramolecular electron transfer between the clusters. In the present study, evidence supporting this idea was obtained by demonstrating that it is possible to alter the rate of substrate reduction by perturbing the polypeptide environment between the P cluster and FeMo-cofactor without necessarily disrupting the metallocluster polypeptide environments or altering component-protein interaction.

Amino Acid Sequence↗
PubMed1995

Nitrogenase structure and function: a biochemical-genetic perspective.

Biological nitrogen fixation is catalyzed by nitrogenase, an enzyme composed of two component proteins called the Fe protein and the MoFe protein. During catalysis, electrons are delivered one at a time from the Fe protein to the MoFe protein in a process involving component-protein association and dissociation and hydrolysis of at least two MgATP for each electron transfer. The Fe protein contains the sites for MgATP binding and hydrolysis, whereas the site for substrate binding and reduction is located on the MoFe protein. Among the important aspects of nitrogenase enzymology discussed here are (a) the structures of the metal centers that participate in electron transfer, (b) the organization of the metalloclusters within the polypeptides and their contributions to substrate binding and electron transfer, (c) the nature of the dynamic interactions between the two component proteins that lead to nucleotide hydrolysis and intermolecular electron transfer, (d) the mechanism by which the multiple electrons necessary for substrate reduction are distributed within the MoFe protein, (e) the nature of the intramolecular electron path within the MoFe protein, and (f) where and how substrate and various inhibitors become bound to the substrate-reduction site. This chapter summarizes biochemical-genetic strategies used to address these questions and discussed them in the context of the recently proposed three-dimensional models for both the Fe protein and MoFe protein from Azotobacter vinelandii.

Metalloproteins↗