Structural study of the X-ray-induced enzymatic reduction of molecular oxygen to water bySteccherinum murashkinskyilaccase: insights into the reaction mechanism

2017 ◽  
Vol 73 (5) ◽  
pp. 388-401 ◽  
Author(s):  
K. M. Polyakov ◽  
S. Gavryushov ◽  
S. Ivanova ◽  
T. V. Fedorova ◽  
O. A. Glazunova ◽  
...  
Keyword(s):  
Molecular Oxygen ◽  
Catalytic Reduction ◽  
Copper Ions ◽  
Absorbed Dose ◽  
Large Family ◽  
Data Sets ◽  
X Ray ◽  
Substrate Complex ◽  
Enzymatic Reduction ◽  
Wide Range

The laccase fromSteccherinum murashkinskyiis a member of the large family of multicopper oxidases that catalyze the oxidation of a wide range of organic and inorganic substrates, accompanied by the reduction of dioxygen to water. The reducing properties of X-ray radiation and the high quality of the laccase crystals allow the study of the catalytic reduction of dioxygen to water directly in a crystal. A series of diffraction data sets with increasing absorbed radiation dose were collected from a single crystal ofSteccherinum murashkinskyilaccase at 1.35 Å resolution. Changes in the active-site structure associated with the reduction of molecular oxygen to water on increasing the absorbed dose of ionizing radiation were detected. The structures in the series are mixtures of different states of the enzyme–substrate complex. Nevertheless, it was possible to interpret these structures as complexes of various oxygen ligands with copper ions in different oxidation states. The results allowed the mechanism of oxygen reduction catalyzed by laccases to be refined.

The subatomic resolution study of laccase inhibition by chloride and fluoride anions using single-crystal serial crystallography: insights into the enzymatic reaction mechanism

2019 ◽  
Vol 75 (9) ◽  
pp. 804-816 ◽  
Author(s):  
Konstantin M. Polyakov ◽  
Sergei Gavryushov ◽  
Tatiana V. Fedorova ◽  
Olga A. Glazunova ◽  
Alexander N. Popov
Keyword(s):  
Molecular Oxygen ◽  
Enzymatic Reaction ◽  
Copper Ion ◽  
Data Sets ◽  
X Ray Diffraction ◽  
Fluoride Ions ◽  
X Ray ◽  
Mechanism Of Inhibition ◽  
Wide Range ◽  
Oxygen Ligand

Laccases are enzymes that catalyze the oxidation of a wide range of organic and inorganic substrates accompanied by the reduction of molecular oxygen to water. Here, a subatomic resolution X-ray crystallographic study of the mechanism of inhibition of the laccase from the basidiomycete fungus Steccherinum murashkinskyi by chloride and fluoride ions is presented. Three series of X-ray diffraction data sets were collected with increasing doses of absorbed X-ray radiation from a native S. murashkinskyi laccase crystal and from crystals of complexes of the laccase with chloride and fluoride ions. The data for the native laccase crystal confirmed the previously deduced enzymatic mechanism of molecular oxygen reduction. The structures of the complexes allowed the localization of chloride and fluoride ions in the channel near the T2 copper ion. These ions replace the oxygen ligand of the T2 copper ion in this channel and can play the role of this ligand in the enzymatic reaction. As follows from analysis of the structures from the increasing dose series, the inhibition of laccases by chloride and fluoride anions can be explained by the fact that the binding of these negatively charged ions at the position of the oxygen ligand of the T2 copper ion impedes the reduction of the T2 copper ion.

Structural study of the X-ray-induced enzymatic reaction of octahaem cytochromecnitrite reductase

2015 ◽  
Vol 71 (5) ◽  
pp. 1087-1094 ◽  
Author(s):  
A. A. Trofimov ◽  
K. M. Polyakov ◽  
V. A. Lazarenko ◽  
A. N. Popov ◽  
T. V. Tikhonova ◽  
...  
Keyword(s):  
Nitrite Reductase ◽  
Catalytic Reaction ◽  
Active Sites ◽  
Enzymatic Reaction ◽  
Absorbed Dose ◽  
Free Form ◽  
Data Sets ◽  
X Rays ◽  
X Ray ◽  
Site Water

Octahaem cytochromecnitrite reductase from the bacteriumThioalkalivibrio nitratireducenscatalyzes the reduction of nitrite to ammonium and of sulfite to sulfide. The reducing properties of X-ray radiation and the high quality of the enzyme crystals allow study of the catalytic reaction of cytochromecnitrite reductase directly in a crystal of the enzyme, with the reaction being induced by X-rays. Series of diffraction data sets with increasing absorbed dose were collected from crystals of the free form of the enzyme and its complexes with nitrite and sulfite. The corresponding structures revealed gradual changes associated with the reduction of the catalytic haems by X-rays. In the case of the nitrite complex the conversion of the nitrite ions bound in the active sites to NO species was observed, which is the beginning of the catalytic reaction. For the free form, an increase in the distance between the oxygen ligand bound to the catalytic haem and the iron ion of the haem took place. In the case of the sulfite complex no enzymatic reaction was detected, but there were changes in the arrangement of the active-site water molecules that were presumably associated with a change in the protonation state of the sulfite ions.

scholarly journals Poisson errors and adaptive rebinning in X-ray powder diffraction data

2018 ◽  
Vol 33 (4) ◽  
pp. 266-269 ◽  
Author(s):  
Marcus H. Mendenhall
Keyword(s):  
Powder Diffraction ◽  
Diffraction Data ◽  
Data Sets ◽  
Powder Diffraction Data ◽  
Data Set ◽  
X Ray ◽  
Short Summary ◽  
Correct Assignment ◽  
Wide Range ◽  
Diffraction Patterns

This work provides a short summary of techniques for formally-correct handling of statistical uncertainties in Poisson-statistics dominated data, with emphasis on X-ray powder diffraction patterns. Correct assignment of uncertainties for low counts is documented. Further, we describe a technique for adaptively rebinning such data sets to provide more uniform statistics across a pattern with a wide range of count rates, from a few (or no) counts in a background bin to on-peak regions with many counts. This permits better plotting of data and analysis of a smaller number of points in a fitting package, without significant degradation of the information content of the data set. Examples of the effect of this on a diffraction data set are given.

scholarly journals Promotional Effect of Cerium and/or Zirconium Doping on Cu/ZSM-5 Catalysts for Selective Catalytic Reduction of NO by NH3

Catalysts ◽  
2018 ◽  
Vol 8 (8) ◽  
pp. 306 ◽  
Author(s):  
Ye Liu ◽  
Chonglin Song ◽  
Gang Lv ◽  
Chenyang Fan ◽  
Xiaodong Li
Keyword(s):  
Selective Catalytic Reduction ◽  
Photoelectron Spectroscopy ◽  
Catalytic Reduction ◽  
Copper Ions ◽  
N2o Formation ◽  
Copper Species ◽  
X Ray ◽  
Promotional Effect ◽  
Zirconium Doping ◽  
Temperature Programmed

The cerium and/or zirconium-doped Cu/ZSM-5 catalysts (CuCexZr1−xOy/ZSM-5) were prepared by ion exchange and characterized by X-ray diffraction (XRD), transmission electron microscopy (TEM), X-ray photoelectron spectroscopy (XPS), temperature-programmed reduction by hydrogen (H2-TPR). Activities of the catalysts obtained on the selective catalytic reduction (SCR) of nitric oxide (NO) by ammonia were measured using temperature programmed reactions. Among all the catalysts tested, the CuCe0.75Zr0.25Oy/ZSM-5 catalyst presented the highest catalytic activity for the removal of NO, corresponding to the broadest active window of 175–468 °C. The cerium and zirconium addition enhanced the activity of catalysts, and the cerium-rich catalysts exhibited more excellent SCR activities as compared to the zirconium-rich catalysts. XRD and TEM results indicated that zirconium additions improved the copper dispersion and prevented copper crystallization. According to XPS and H2-TPR analysis, copper species were enriched on the ZSM-5 grain surfaces, and part of the copper ions were incorporated into the zirconium and/or cerium lattice. The strong interaction between copper species and cerium/zirconium improved the redox abilities of catalysts. Furthermore, the introduction of zirconium abates N2O formation in the tested temperature range.

Quantitative Electron Microprobe Analysis Of Semiconductor Materials: An Evaluation Of Accuracy

1999 ◽  
Vol 5 (S2) ◽  
pp. 74-75
Author(s):  
P.K. Carpenter
Keyword(s):  
Ternary Systems ◽  
Data Sets ◽  
Silicate Mineral ◽  
Data Set ◽  
X Ray ◽  
Quantitative Microanalysis ◽  
Wide Range ◽  
Analytical Accuracy ◽  
Unknown Composition ◽  
Counting Statistics

Both precision and accuracy are central to quantitative microanalysis. While precision may be evaluated from x-ray counting statistics and replicate measurement, the determination of analytical accuracy requires well characterized standards of which there are few that span a wide range of compositions in binary and ternary systems. The accuracy of silicate mineral analysis has been previously studied via measurement of α factors at multiple accelerating potential and the subsequent evaluation of correction algorithms and mass absorption coefficient (mac) data sets. This approach has been extended in this study to the In2O3-Ga2O3 and HgTe-CdTe systems.Single crystals of ln2O3, Ga2O3, and an InGa-oxide of unknown composition were used to evaluate accuracy in the In2O3-Ga2O3 binary, using the GaKα, GaLα, and InLα x-ray lines, with WDS measurements performed at 15, 20, and 25KV relative to the ln2O3 and Ga2O3 standards (see Table I). The Ga Kα line exhibits minimal absorption, has no fluorescence correction in this system and is not critically dependent on the correction algorithm or mac data set used.

scholarly journals Crystallization and preliminary X-ray crystallographic analysis of the catalytic domain of membrane type 1 matrix metalloproteinase

2014 ◽  
Vol 70 (2) ◽  
pp. 232-235 ◽  
Author(s):  
Hideaki Ogata ◽  
Elena Decaneto ◽  
Moran Grossman ◽  
Martina Havenith ◽  
Irit Sagi ◽  
...  
Keyword(s):  
Matrix Metalloproteinase ◽  
Catalytic Domain ◽  
Large Family ◽  
Crystallographic Analysis ◽  
Diffusion Method ◽  
Data Sets ◽  
X Ray ◽  
Cell Parameters ◽  
Membrane Type

Membrane type 1 matrix metalloproteinase (MT1-MMP) belongs to the large family of zinc-dependent endopeptidases termed MMPs that are located in the extracellular matrix. MT1-MMP was crystallized at 277 K using the vapour-diffusion method with PEG as a precipitating agent. Data sets for MT1-MMP were collected to 2.24 Å resolution at 100 K. The crystals belonged to space groupP43212, with unit-cell parametersa= 62.99,c= 122.60 Å. The crystal contained one molecule per asymmetric unit, with a Matthews coefficient (VM) of 2.90 Å3 Da−1; the solvent content is estimated to be 57.6%.

scholarly journals Quantum Mechanical Study of Protonation of Oxygen Ligands in the Laccase Active Site Based on X-Ray Structures of Subatomic Resolution

2022 ◽  
Author(s):  
Sergei Gavryushov ◽  
Nikolay Kuzmich ◽  
Konstantin Polyakov
Keyword(s):  
Oxygen Reduction ◽  
Molecular Oxygen ◽  
High Precision ◽  
Active Site ◽  
Quantum Mechanical ◽  
Quantum Mechanical Calculations ◽  
Stable States ◽  
Hydrogen Atoms ◽  
X Ray ◽  
Wide Range

Laccases are enzymes catalyzing oxidation of a wide range of organic and inorganic substrates accompanied by molecular oxygen reduction to water. Previously studies of oxygen reduction by laccases have recently been reported. They were based on single-crystal serial X-ray crystallography with increasing absorption doses at subatomic resolution, As a result, coordinates of all non-hydrogen atoms of the active site have been determined with high precision for both oxidized and reduced states of the enzyme. Those data can be used to clarify the mechanism of molecular oxygen reduction by laccases. However, the X-ray data lack information about protonation states of the oxygen ligands involved. Applying quantum mechanical calculations, in the present work protonation of oxygen ligands in the active site of laccase was determined for both reduced and oxidized states of the enzyme (the stable states observed in experiments at reduction of molecular oxygen in laccase). The high precision of X-ray-determined atom coordinates allowed us to simplify preliminary calculations of molecular mechanics for models used in the quantum mechanical calculations.

Gas-sensitive biological crystals processed in pressurized oxygen and krypton atmospheres: deciphering gas channels in proteins using a novel `soak-and-freeze' methodology

2016 ◽  
Vol 49 (5) ◽  
pp. 1478-1487 ◽  
Author(s):  
Bénédicte Lafumat ◽  
Christoph Mueller-Dieckmann ◽  
Gordon Leonard ◽  
Nathalie Colloc'h ◽  
Thierry Prangé ◽  
...  
Keyword(s):  
High Pressure ◽  
Molecular Oxygen ◽  
Gas Pressure ◽  
Biological Molecules ◽  
Urate Oxidase ◽  
Biological Processes ◽  
X Ray ◽  
Pressure Cell ◽  
Sufficient Detail ◽  
Wide Range

Molecular oxygen (O2) is a key player in many fundamental biological processes. However, the combination of the labile nature and poor affinity of O2 often makes this substrate difficult to introduce into crystals at sufficient concentrations to enable protein/O2 interactions to be deciphered in sufficient detail. To overcome this problem, a gas pressure cell has been developed specifically for the `soak-and-freeze' preparation of crystals of O2-dependent biological molecules. The `soak-and-freeze' method uses high pressure to introduce oxygen molecules or krypton atoms (O2 mimics) into crystals which, still under high pressure, are then cryocooled for X-ray data collection. Here, a proof of principle of the gas pressure cell and the methodology developed is demonstrated with crystals of enzymes (lysozyme, thermolysin and urate oxidase) that are known to absorb and bind molecular oxygen and/or krypton. The successful results of these experiments lead to the suggestion that the soak-and-freeze method could be extended to studies involving a wide range of gases of biological, medical and/or environmental interest, including carbon monoxide, ethylene, methane and many others.

Elucidation of the crystal structure ofCoriolopsis caperatalaccase: restoration of the structure and activity of the native enzyme from the T2-depleted form by copper ions

2015 ◽  
Vol 71 (4) ◽  
pp. 854-861 ◽  
Author(s):  
Olga A. Glazunova ◽  
Konstantin M. Polyakov ◽  
Tatyana V. Fedorova ◽  
Pavel V. Dorovatovskii ◽  
Olga V. Koroleva
Keyword(s):  
Copper Ions ◽  
Large Family ◽  
Native Enzyme ◽  
Multicopper Oxidases ◽  
Wide Range ◽  
Inorganic Substrates ◽  
Structure And Activity ◽  
Copper Centre ◽  
High Redox Potential

Laccases are members of a large family of multicopper oxidases that catalyze the oxidation of a wide range of organic and inorganic substrates accompanied by the reduction of dioxygen to water. A new laccase was isolated from the basidiomyceteCoriolopsis caperatastrain 0677 and its amino-acid sequence was determined. According to its physicochemical properties and spectroscopic features, the laccase fromC. caperatais a high redox-potential blue laccase. Attempts to crystallize the native enzyme were unsuccessful. The copper type 2-depleted (T2D) laccase was prepared and crystallized. The structure of T2D laccase fromC. caperatawas solved at 1.6 Å resolution, and attempts to reconstruct the T2 copper centre were performed using Cu+and Cu2+ions. The structure of T2D+Cu+laccase was solved at 1.89 Å resolution. It was shown that the T2D+Cu+laccase structure contained four copper ions in the active site. Reconstruction could not be achieved when the T2D laccase crystals were treated with CuSO4.

scholarly journals Photoevaporation versus core-powered mass-loss: model comparison with the 3D radius gap

2021 ◽  
Author(s):  
James G Rogers ◽  
Akash Gupta ◽  
James E Owen ◽  
Hilke E Schlichting
Keyword(s):  
Mass Loss ◽  
Model Comparison ◽  
Hypothesis Test ◽  
Space Mission ◽  
Data Sets ◽  
X Ray ◽  
Wide Range ◽  
Cent Level ◽  
Loss Models ◽  
Atmospheric Mass

Abstract The EUV/X-ray photoevaporation and core-powered mass-loss models are both capable of reproducing the bimodality in the sizes of small, close-in exoplanets observed by the Kepler space mission, often referred to as the ‘radius gap’. However, it is unclear which of these two mechanisms dominates the atmospheric mass-loss which is likely sculpting the radius gap. In this work, we propose a new method of differentiating between the two models, which relies on analysing the radius gap in 3D parameter space. Using models for both mechanisms, and by performing synthetic transit surveys we predict the size and characteristics of a survey capable of discriminating between the two models. We find that a survey of ≳ 5000 planets, with a wide range in stellar mass and measurement uncertainties at a $\lesssim 5{{\ \rm per\ cent}}$ level is sufficient. Our methodology is robust against moderate false positive contamination of $\lesssim 10{{\ \rm per\ cent}}$. We perform our analysis on two surveys (which do not satisfy our requirements): the California Kepler Survey and the Gaia-Kepler Survey and find, unsurprisingly, that both data-sets are consistent with either model. We propose a hypothesis test to be performed on future surveys which can robustly ascertain which of the two mechanisms formed the radius gap, provided one dominates over the other.

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