Analysis of the Stability of Mutant Lysozymes at Position 15 Using X-Ray Crystallography

1997 ◽  
Vol 122 (3) ◽  
pp. 512-517 ◽  
Author(s):  
T. Ohmura ◽  
T. Ueda ◽  
H. Motoshima ◽  
T. Tamura ◽  
T. Imoto
Keyword(s):  
X Ray ◽  
X Ray Crystallography ◽  
The Stability

scholarly journals Structure of human RNA Polymerase III

2020 ◽  
Author(s):  
Ewan Phillip Ramsay ◽  
Guillermo Abascal-Palacios ◽  
Julia L. Daiß ◽  
Helen King ◽  
Jerome Gouge ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Iii ◽  
Cellular Growth ◽  
Loss Of Function ◽  
Critical Determinant ◽  
Saxs Data ◽  
X Ray ◽  
X Ray Crystallography ◽  
The Stability ◽  
Pol Iii

ABSTRACTIn eukaryotes, RNA Polymerase (Pol) III is the enzyme specialised for the transcription of the entire pool of tRNAs and several other short, essential, untranslated RNAs. Pol III is a critical determinant of cellular growth and lifespan across the eukaryotic kingdom. Upregulation of Pol III transcription is often observed in cancer cells and causative Pol III mutations have been described in patients affected by severe neurodevelopmental disorders and hypersensitivity to viral infection.Harnessing CRISPR-Cas9 genome editing in HeLa cells, we isolated endogenous human Pol III and obtained a cryo-EM reconstruction at 4.0 Å. The structure of human Pol III allowed us to map the reported genetic mutations and rationalise them. Mutations causing neurodevelopmental defects cluster in hotspots that affect the stability and/or biogenesis of Pol III, thereby resulting in loss-of-function of the enzyme. Mutations affecting viral sensing are located in the periphery of the enzyme in proximity to DNA binding regions, suggesting an impairment of Pol III cytosolic viral DNA-sensing activity.Furthermore, integrating x-ray crystallography and SAXS data, we describe the structure of the RPC5 C-terminal extension, which is absent in lower eukaryotes and not visible in our EM map. Surprisingly, experiments in living cells highlight a role for the RPC5 C-terminal extension in the correct assembly and stability of the human Pol III enzyme, thus suggesting an added layer of regulation during the biogenesis of Pol III in higher eukaryotes.

scholarly journals Structural insights into the small G-protein Arl13B and implications for Joubert syndrome

2013 ◽  
Vol 457 (2) ◽  
pp. 301-311 ◽  
Author(s):  
Mandy Miertzschke ◽  
Carolin Koerner ◽  
Michael Spoerner ◽  
Alfred Wittinghofer
Keyword(s):  
High Resolution ◽  
G Protein ◽  
Joubert Syndrome ◽  
Active Conformation ◽  
Loss Of Function ◽  
X Ray ◽  
X Ray Crystallography ◽  
Small G Protein ◽  
The Stability ◽  
Structural Insights

Using Arl13B from Chlamydomonas as a model, we show by high resolution X-ray crystallography and biochemical approaches that mutations in patients with Joubert syndrome might lead to loss of function by specifically affecting the stability of the active conformation of Arl13B.

scholarly journals Cevipabulin-tubulin complex reveals a novel agent binding site on α-tubulin with tubulin degradation effect

2021 ◽  
Vol 7 (21) ◽  
pp. eabg4168
Author(s):  
Jianhong Yang ◽  
Yamei Yu ◽  
Yong Li ◽  
Wei Yan ◽  
Haoyu Ye ◽  
...  
Keyword(s):  
Binding Site ◽  
Binding Sites ◽  
X Ray ◽  
X Ray Crystallography ◽  
The Stability ◽  
New Generation ◽  
Antimicrotubule Drugs ◽  
Degradation Effect ◽  
Microtubule Stabilizing Agent ◽  
Shed Light

Microtubules, composed of αβ-tubulin heterodimers, have remained popular anticancer targets for decades. Six known binding sites on tubulin dimers have been identified thus far, with five sites on β-tubulin and only one site on α-tubulin, hinting that compounds binding to α-tubulin are less well characterized. Cevipabulin, a microtubule-active antitumor clinical candidate, is widely accepted as a microtubule-stabilizing agent by binding to the vinblastine site. Our x-ray crystallography study reveals that, in addition to binding to the vinblastine site, cevipabulin also binds to a new site on α-tubulin. We find that cevipabulin at this site pushes the αT5 loop outward, making the nonexchangeable GTP exchangeable, which reduces the stability of tubulin, leading to its destabilization and degradation. Our results confirm the existence of a new agent binding site on α-tubulin and shed light on the development of tubulin degraders as a new generation of antimicrotubule drugs targeting this novel site.

scholarly journals Using crystallography tools to improve vaccine formulations

IUCrJ ◽  
2021 ◽  
Vol 9 (1) ◽  
Author(s):  
Márcia Carvalho de Abreu Fantini ◽  
Cristiano Luis Pinto Oliveira ◽  
José Luiz de Souza Lopes ◽  
Tereza da Silva Martins ◽  
Milena Apetito Akamatsu ◽  
...  
Keyword(s):  
Oral Vaccine ◽  
Porous Silica ◽  
Protein Antigens ◽  
X Ray ◽  
X Ray Crystallography ◽  
X Ray Scattering ◽  
Transmission Electron ◽  
X Ray Imaging ◽  
The Stability ◽  
Phase Contrast Tomography

This article summarizes developments attained in oral vaccine formulations based on the encapsulation of antigen proteins inside porous silica matrices. These vaccine vehicles show great efficacy in protecting the proteins from the harsh acidic stomach medium, allowing the Peyer's patches in the small intestine to be reached and consequently enhancing immunity. Focusing on the pioneering research conducted at the Butantan Institute in Brazil, the optimization of the antigen encapsulation yield is reported, as well as their distribution inside the meso- and macroporous network of the porous silica. As the development of vaccines requires proper inclusion of antigens in the antibody cells, X-ray crystallography is one of the most commonly used techniques to unveil the structure of antibody-combining sites with protein antigens. Thus structural characterization and modelling of pure antigen structures, showing different dimensions, as well as their complexes, such as silica with encapsulated hepatitis B virus-like particles and diphtheria anatoxin, were performed using small-angle X-ray scattering, X-ray absorption spectroscopy, X-ray phase contrast tomography, and neutron and X-ray imaging. By combining crystallography with dynamic light scattering and transmission electron microscopy, a clearer picture of the proposed vaccine complexes is shown. Additionally, the stability of the immunogenic complex at different pH values and temperatures was checked and the efficacy of the proposed oral immunogenic complex was demonstrated. The latter was obtained by comparing the antibodies in mice with variable high and low antibody responses.

Mercury(II) and Lead(II) Complexes of 1-Oxa-7-thia-4,10-diazacyclododecane ([12]aneN2OS)

1999 ◽  
Vol 52 (1) ◽  
pp. 1 ◽  
Author(s):  
Trevor W. Hambley ◽  
Shahara Afshar ◽  
Sebastian T. Marcus ◽  
Lawrence R. Gahan
Keyword(s):  
Stability Constants ◽  
Macrocyclic Ligand ◽  
Protonation Constants ◽  
Secondary Amines ◽  
Monoclinic Space Group ◽  
Space Group ◽  
X Ray ◽  
X Ray Crystallography ◽  
The Stability ◽  
Lead Complex

The mixed donor 12-membered macrocyclic ligand 1-oxa-7-thia-4,10-diazacyclododecane ([12]aneN2OS) has been synthesized and the mercury(II) and lead(II) complexes, [Hg([12]aneN2OS)(NO3)2] and [Pb([12]aneN2OS)(NO3)2], have been isolated and characterized by X-ray crystallography. Crystals of the mercury complex are monoclinic, space group P 21/c, a 9·576(2), b 10·757(2), c 14·789(4) Å, β 93·58(2)°, whilst crystals of the lead complex are monoclinic, space group P 21/n, a 19·490(7), b 8·010(2), c 19·576(6) Å, β 109·90(2)°. The protonation constants and stability constants have been determined potentiometrically in aqueous solution. The protonation constants for [12]aneN2OS (log KHL 9·13; log K H2L 6·85) appear typical for secondary amines in similar trans-substituted 12-membered macrocycles. The magnitudes of the stability constants (HgII, log KHgL 10·5; PbII, log KPbL 6·6) are consistent with trends observed previously for macrocyclic ligands as secondary amine donors are replaced with oxygen and thioether donors.

Syntheses, crystal structures and phosphorescence properties of cyclometalated iridium(III) bis(pyridylbenzaldehyde) complexes with dithiolate ligands

2017 ◽  
Vol 72 (12) ◽  
pp. 941-946 ◽  
Author(s):  
Xue-Mei Wang ◽  
Jia-Yan Qiang ◽  
Ai-Quan Jia ◽  
Bihai Tong ◽  
Qian-Feng Zhang
Keyword(s):  
Crystal Structures ◽  
Emission Spectra ◽  
Transition Metal Ions ◽  
X Ray ◽  
X Ray Crystallography ◽  
H Nmr ◽  
Water Solvent ◽  
The Stability ◽  
C Nmr ◽  
Composition Of Complexes

AbstractThe synthesis of three neutral bis-cyclometalated iridium(III) complexes [Ir(pba)2(S^S)] (pbaH=4-(2-pyridyl)benzaldehyde; S^S=Et2NCS2− (1), iPrOCS2− (2), (nPrO)2PS2− (3)) from [{Ir(μ-Cl)(pba)2}2] and the corresponding sodium or potassium dithiolates in methanol-dichloromethane is described. The composition of complexes 1–3 is discussed on the basis of 1H NMR, 13C NMR, IR, and mass spectroscopy, and the crystal structures of 1 and 3 were determined by X-ray crystallography. The absorption and emission spectra show that the [Ir(pba)2(S^S)] complexes may be effective candidates as green-emitting phosphorescent materials. The stability of the three cyclometalated iridium(III) complexes towards different transition metal ions was also investigated in acetonitrile-water solvent.

Novel Synthesis, Molecular Structure, and Theoretical Studies of Dispiro Compounds via Pseudo-eight-component Reaction

2014 ◽  
Vol 67 (11) ◽  
pp. 1656 ◽  
Author(s):  
Nourallah Hazeri ◽  
Mojtaba Lashkari ◽  
Santiago García-Granda ◽  
Laura Torre-Fernández
Keyword(s):  
Diels Alder ◽  
Quantum Mechanical ◽  
Theoretical Studies ◽  
Diastereoselective Synthesis ◽  
One Pot ◽  
X Ray ◽  
Complex Products ◽  
X Ray Crystallography ◽  
Novel Synthesis ◽  
The Stability

We have developed a diastereoselective synthesis of dispiro compounds through a one-pot domino pseudo-eight-component reaction of amines, aldehydes, and Meldrum’s acid. This method resulted in the generation of complex products with four stereocentres and involves formation of 10 new bonds. Quantum mechanical calculations were undertaken in order to determine the stability of the eight diastereomer structures of compound 4a. The crystal structure of 4k was determined by X-ray crystallography. Reaction mechanism can proceed through Knoevenagel, Aldol condensations, Diels-Alder cycloaddition, and Michael addition.

scholarly journals Analyses of the stability and function of three surface mutants (R82C, K69H, and L32R) of the gene V protein from Ff phage by X-ray crystallography

1997 ◽  
Vol 6 (4) ◽  
pp. 771-780 ◽  
Author(s):  
Shaoyu Su ◽  
Yi-Gui Gao ◽  
Hong Zhang ◽  
Thomas C. Terwilliger ◽  
ANDREW H.-J. Wang
Keyword(s):  
V Protein ◽  
X Ray ◽  
X Ray Crystallography ◽  
The Stability ◽  
And Function

Difference Fourier Analysis of Glucose Embedded and Frozen Hydrated Purple Membrane

1982 ◽  
Vol 40 ◽  
pp. 74-75
Author(s):  
Jules S. Jaffe ◽  
Robert M. Glaeser
Keyword(s):  
Purple Membrane ◽  
Data Set ◽  
High Resolution Data ◽  
X Ray ◽  
X Ray Crystallography ◽  
Fourier Techniques ◽  
Versus Protein ◽  
The Difference ◽  
Difference Fourier ◽  
Ideal Method

Although difference Fourier techniques are standard in X-ray crystallography it has only been very recently that electron crystallographers have been able to take advantage of this method. We have combined a high resolution data set for frozen glucose embedded Purple Membrane (PM) with a data set collected from PM prepared in the frozen hydrated state in order to visualize any differences in structure due to the different methods of preparation. The increased contrast between protein-ice versus protein-glucose may prove to be an advantage of the frozen hydrated technique for visualizing those parts of bacteriorhodopsin that are embedded in glucose. In addition, surface groups of the protein may be disordered in glucose and ordered in the frozen state. The sensitivity of the difference Fourier technique to small changes in structure provides an ideal method for testing this hypothesis.

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