Understanding the pH-Dependent Reaction Mechanism of a Glycoside Hydrolase Using High-Resolution X-ray and Neutron Crystallography

ACS Catalysis ◽  
2018 ◽  
Vol 8 (9) ◽  
pp. 8058-8069 ◽  
Author(s):  
Zhihong Li ◽  
Xiaoshuai Zhang ◽  
Qingqing Wang ◽  
Chunran Li ◽  
Nianying Zhang ◽  
...  
Keyword(s):  
Reaction Mechanism ◽  
High Resolution ◽  
Glycoside Hydrolase ◽  
X Ray ◽  
Ph Dependent ◽  
Neutron Crystallography

scholarly journals Catalytically important damage-free structures of a copper nitrite reductase obtained by femtosecond X-ray laser and room-temperature neutron crystallography

IUCrJ ◽  
2019 ◽  
Vol 6 (4) ◽  
pp. 761-772 ◽  
Author(s):  
Thomas P. Halsted ◽  
Keitaro Yamashita ◽  
Chai C. Gopalasingam ◽  
Rajesh T. Shenoy ◽  
Kunio Hirata ◽  
...  
Keyword(s):  
High Resolution ◽  
Resting State ◽  
Structural Data ◽  
Redox System ◽  
X Rays ◽  
X Ray ◽  
Nitrite Reductases ◽  
First Time ◽  
Better Than ◽  
Neutron Crystallography

Copper-containing nitrite reductases (CuNiRs) that convert NO2−to NO via a CuCAT–His–Cys–CuETproton-coupled redox system are of central importance in nitrogen-based energy metabolism. These metalloenzymes, like all redox enzymes, are very susceptible to radiation damage from the intense synchrotron-radiation X-rays that are used to obtain structures at high resolution. Understanding the chemistry that underpins the enzyme mechanisms in these systems requires resolutions of better than 2 Å. Here, for the first time, the damage-free structure of the resting state of one of the most studied CuNiRs was obtained by combining X-ray free-electron laser (XFEL) and neutron crystallography. This represents the first direct comparison of neutron and XFEL structural data for any protein. In addition, damage-free structures of the reduced and nitrite-bound forms have been obtained to high resolution from cryogenically maintained crystals by XFEL crystallography. It is demonstrated that AspCATand HisCATare deprotonated in the resting state of CuNiRs at pH values close to the optimum for activity. A bridging neutral water (D2O) is positioned with one deuteron directed towards AspCAT Oδ1and one towards HisCAT N∊2. The catalytic T2Cu-ligated water (W1) can clearly be modelled as a neutral D2O molecule as opposed to D3O+or OD−, which have previously been suggested as possible alternatives. The bridging water restricts the movement of the unprotonated AspCATand is too distant to form a hydrogen bond to the O atom of the bound nitrite that interacts with AspCAT. Upon the binding of NO2−a proton is transferred from the bridging water to the Oδ2atom of AspCAT, prompting electron transfer from T1Cu to T2Cu and reducing the catalytic redox centre. This triggers the transfer of a proton from AspCATto the bound nitrite, enabling the reaction to proceed.

scholarly journals Structure of HIV-1 protease in complex with potent inhibitor KNI-272 determined by high-resolution X-ray and neutron crystallography

2009 ◽  
Vol 106 (12) ◽  
pp. 4641-4646 ◽  
Author(s):  
M. Adachi ◽  
T. Ohhara ◽  
K. Kurihara ◽  
T. Tamada ◽  
E. Honjo ◽  
...  
Keyword(s):  
High Resolution ◽  
Potent Inhibitor ◽  
X Ray ◽  
Hiv 1 ◽  
Neutron Crystallography

scholarly journals Studying the Role of a Single Mutation of a Family 11 Glycoside Hydrolase Using High-Resolution X-ray Crystallography

2020 ◽  
Vol 39 (6) ◽  
pp. 671-680
Author(s):  
Zhihong Li ◽  
Xiaoshuai Zhang ◽  
Chunran Li ◽  
Andrey Kovalevsky ◽  
Qun Wan
Keyword(s):  
High Resolution ◽  
Glycoside Hydrolase ◽  
Single Mutation ◽  
X Ray ◽  
X Ray Crystallography

Electron microscopy of rabbit FC crystals

1983 ◽  
Vol 41 ◽  
pp. 730-731
Author(s):  
Robert A. Grant ◽  
Laura L. Degn ◽  
Wah Chiu ◽  
John Robinson
Keyword(s):  
Electron Microscopy ◽  
High Resolution ◽  
High Resolution Electron Microscopy ◽  
Molecular Replacement ◽  
X Ray ◽  
X Ray Crystallography ◽  
Resolution Electron ◽  
Replacement Technique ◽  
Hydrated Crystal ◽  
Molecular Structure Analysis

Proteolytic digestion of the immunoglobulin IgG with papain cleaves the molecule into an antigen binding fragment, Fab, and a compliment binding fragment, Fc. Structures of intact immunoglobulin, Fab and Fc from various sources have been solved by X-ray crystallography. Rabbit Fc can be crystallized as thin platelets suitable for high resolution electron microscopy. The structure of rabbit Fc can be expected to be similar to the known structure of human Fc, making it an ideal specimen for comparing the X-ray and electron crystallographic techniques and for the application of the molecular replacement technique to electron crystallography. Thin protein crystals embedded in ice diffract to high resolution. A low resolution image of a frozen, hydrated crystal can be expected to have a better contrast than a glucose embedded crystal due to the larger density difference between protein and ice compared to protein and glucose. For these reasons we are using an ice embedding technique to prepare the rabbit Fc crystals for molecular structure analysis by electron microscopy.

High Resolution Study of Polytypism: Application to SiC and Au3 Mn

1982 ◽  
Vol 40 ◽  
pp. 540-541
Author(s):  
G. Van Tendeloo ◽  
J. Van Landuyt ◽  
S. Amelinckx
Keyword(s):  
Electron Microscopy ◽  
High Resolution ◽  
Stacking Sequence ◽  
X Ray ◽  
Powerful Technique ◽  
Resolution Study ◽  
The Ideal

Polytypism has been studied for a number of years and a wide variety of stacking sequences has been detected and analysed. SiC is the prototype material in this respect; see e.g. Electron microscopy under high resolution conditions when combined with x-ray measurements is a very powerful technique to elucidate the correct stacking sequence or to study polytype transformations and deviations from the ideal stacking sequence.

A High Resolution Study of G.P. Zones in Aluminum - 4.2 at % Silver

1982 ◽  
Vol 40 ◽  
pp. 722-723 ◽  
Author(s):  
R. Gronsky
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
High Resolution ◽  
Computer Simulations ◽  
Structural Characteristics ◽  
X Ray Diffraction ◽  
X Ray ◽  
Transmission Electron ◽  
Resolution Study

The phenomenon of clustering in Al-Ag alloys has been extensively studied since the early work of Guinierl, wherein the pre-precipitation state was characterized as an assembly of spherical, ordered, silver-rich G.P. zones. Subsequent x-ray and TEM investigations yielded results in general agreement with this model. However, serious discrepancies were later revealed by the detailed x-ray diffraction - based computer simulations of Gragg and Cohen, i.e., the silver-rich clusters were instead octahedral in shape and fully disordered, atleast below 170°C. The object of the present investigation is to examine directly the structural characteristics of G.P. zones in Al-Ag by high resolution transmission electron microscopy.

Scanning Electron Microscopy and x-ray analysis of microparticles and early hydration effects

1995 ◽  
Vol 53 ◽  
pp. 692-693
Author(s):  
Yun Lu ◽  
David C. Joy
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
High Resolution ◽  
Morphological Development ◽  
Early Hydration ◽  
X Ray ◽  
Blended Cements ◽  
Powder Particles ◽  
Chemical Wastes ◽  
Scanning Electron

High resolution scanning electron microscopy (SEM) and energy dispersive x-ray analysis (EDXA) were performed to investigate microparticles in blended cements and their hydration products containing sodium-rich chemical wastes. The physical appearance of powder particles and the morphological development at different hydration stages were characterized by using high resolution SEM Hitachi S-900 and by SEM S-800 with a EDX spectrometer. Microparticles were dispersed on the sample holder and glued by 1% palomino solution. Hydrated bulk samples were dehydrated by acetone and mounted on the holder by silver paste. Both fracture surfaces and flat cutting sections of hydrating samples were prepared and examined. Some specimens were coated with an 3 nm thick Au-Pd or Cr layer to provide good conducting surfaces. For high resolution SEM S-900 observations the accelerating voltage of electrons was 1-2 KeV to protect the electron charging. Microchemical analyses were carried out by S800/EDS equipped with a LINK detector of take-off angle =40°.

Tubulin structure at intermediate resolution

1995 ◽  
Vol 53 ◽  
pp. 852-853
Author(s):  
K. H. Downing ◽  
S. G. Wolf ◽  
E. Nogales
Keyword(s):  
High Resolution ◽  
Structural Data ◽  
Therapeutic Drug ◽  
Zinc Ions ◽  
Two Dimensional ◽  
X Ray Diffraction ◽  
X Ray ◽  
Negative Stain ◽  
Functional Mechanisms ◽  
Tubulin Structure

Microtubules are involved in a host of critical cell activities, many of which involve transport of organelles through the cell. Different sets of microtubules appear to form during the cell cycle for different functions. Knowledge of the structure of tubulin will be necessary in order to understand the various functional mechanisms of microtubule assemble, disassembly, and interaction with other molecules, but tubulin has so far resisted crystallization for x-ray diffraction studies. Fortuitously, in the presence of zinc ions, tubulin also forms two-dimensional, crystalline sheets that are ideally suited for study by electron microscopy. We have refined procedures for forming the sheets and preparing them for EM, and have been able to obtain high-resolution structural data that sheds light on the formation and stabilization of microtubules, and even the interaction with a therapeutic drug.Tubulin sheets had been extensively studied in negative stain, demonstrating that the same protofilament structure was formed in the sheets and microtubules. For high resolution studies, we have found that the sheets embedded in either glucose or tannin diffract to around 3 Å.

Nano-probe x-ray analysis and high-resolution imaging on planar defects in high-pressure synthesized infinite-layer superconductor

1994 ◽  
Vol 52 ◽  
pp. 728-729
Author(s):  
Y. Y. Wang ◽  
H. Zhang ◽  
V. P. Dravid ◽  
H. Zhang ◽  
L. D. Marks ◽  
...  
Keyword(s):  
High Pressure ◽  
High Resolution ◽  
Atomic Structure ◽  
Emission Spectra ◽  
High Resolution Electron Microscopy ◽  
Planar Defects ◽  
X Ray ◽  
Infinite Layer ◽  
Resolution Electron ◽  
Resolution Imaging

Azuma et al. observed planar defects in a high pressure synthesized infinitelayer compound (i.e. ACuO2 (A=cation)), which exhibits superconductivity at ~110 K. It was proposed that the defects are cation deficient and that the superconductivity in this material is related to the planar defects. In this report, we present quantitative analysis of the planar defects utilizing nanometer probe xray microanalysis, high resolution electron microscopy, and image simulation to determine the chemical composition and atomic structure of the planar defects. We propose an atomic structure model for the planar defects.Infinite-layer samples with the nominal chemical formula, (Sr1-xCax)yCuO2 (x=0.3; y=0.9,1.0,1.1), were prepared using solid state synthesized low pressure forms of (Sr1-xCax)CuO2 with additions of CuO or (Sr1-xCax)2CuO3, followed by a high pressure treatment.Quantitative x-ray microanalysis, with a 1 nm probe, was performed using a cold field emission gun TEM (Hitachi HF-2000) equipped with an Oxford Pentafet thin-window x-ray detector. The probe was positioned on the planar defects, which has a 0.74 nm width, and x-ray emission spectra from the defects were compared with those obtained from vicinity regions.

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