Low concentration of guanidine hydrochloride induces the formation of an aggregation-prone state in α-urease

2004 ◽  
Vol 82 (2) ◽  
pp. 305-313 ◽  
Author(s):  
F -O McDuff ◽  
A Doucet ◽  
M Beauregard
Keyword(s):  
Tertiary Structure ◽  
Molten Globule ◽  
Guanidine Hydrochloride ◽  
Average Diameter ◽  
Secondary Structures ◽  
Fluorescence Measurements ◽  
Protein Fibrils ◽  
Electron Microscopy Analysis ◽  
Microscopy Analysis ◽  
The Impact

Canavalia ensiformis (jack bean) α-urease is a hexameric protein characterized by a complex denaturation mechanism. In previous papers, we have shown that a hydrophobic 8-anilino-1-naphthalenesulfonic acid (ANSA) binding conformer could be populated in a moderate concentration of denaturant. This state was obtained under conditions that had no detectable impact on its tertiary structure, as indicated by fluorescence measurements. In the present study, we further characterized this ANSA-binding state in an attempt to understand urease behavior. Evidence presented here shows that the presence of ANSA was not required for the generation of the conformer and that its affinity for ANSA came from an increase in hydrophobicity leading to aggregation. Circular dichroism investigation of urease revealed that it had periodical secondary structure content similar to Klebsiella aerogenes urease (secondary structures calculated on the basis of crystallographic data). The impact of 0.9 M guanidine hydrochloride (GuHCl) on soluble urease secondary structures was minimal but is compatible with a slight increase in beta-sheet structures. Such modification may indicates that aggregation involves amyloid-like fibril formation. Electron microscopy analysis of urease in the absence of GuHCl revealed the presence of urease hexamers (round shape 13 nm in diameter). These particles disappeared in the presence of moderate denaturant concentration owing to the formation of aggregates and fibril-like structures. The fibrils obtained in 1.5 M GuHCl had an average diameter of 6.5 nm, suggesting that urease hexamers dissociated into smaller oligomeric forms when forming such fibrils.Key words: protein structure, protein folding, denaturation, aggregation, multimeric proteins, protein fibrils, hydrophobicity, molten globule state.

Material analysis techniques using the ion mill

1996 ◽  
Vol 54 ◽  
pp. 1034-1035
Author(s):  
J. P. Benedict ◽  
R. M. Anderson ◽  
S. J. Klepeis
Keyword(s):  
Polished Surface ◽  
Surface Material ◽  
Analysis Techniques ◽  
Material Analysis ◽  
Optical Inspection ◽  
Electron Microscopy Analysis ◽  
Microscopy Analysis ◽  
Argon Ions ◽  
The Impact ◽  
Scanning Electron

Ion mills equipped with flood guns can perform two important functions in material analysis; they can either remove material or deposit material. The ion mill holder shown in Fig. 1 is used to remove material from the polished surface of a sample for further optical inspection or SEM ( Scanning Electron Microscopy ) analysis. The sample is attached to a pohshing stud type SEM mount and placed in the ion mill holder with the polished surface of the sample pointing straight up, as shown in Fig 2. As the holder is rotating in the ion mill, Argon ions from the flood gun are directed down at the top of the sample. The impact of Argon ions against the surface of the sample causes some of the surface material to leave the sample at a material dependent, nonuniform rate. As a result, the polished surface will begin to develop topography during milling as fast sputtering materials leave behind depressions in the polished surface.

scholarly journals Impact Resistance of Epoxy Composites Reinforced with Amazon Guaruman Fiber: A Brief Report

Polymers ◽  
2021 ◽  
Vol 13 (14) ◽  
pp. 2264
Author(s):  
Raphael H. M. Reis ◽  
Fabio C. Garcia Filho ◽  
Larissa F. Nunes ◽  
Veronica S. Candido ◽  
Alisson C. R. Silva ◽  
...  
Keyword(s):  
Fiber Composite ◽  
Impact Resistance ◽  
Fiber Composites ◽  
Epoxy Composites ◽  
Impact Performance ◽  
Electron Microscopy Analysis ◽  
Ballistic Properties ◽  
Microscopy Analysis ◽  
Izod Impact ◽  
The Impact

Fibers extracted from Amazonian plants that have traditionally been used by local communities to produce simple items such as ropes, nets, and rugs, are now recognized as promising composite reinforcements. This is the case for guaruman (Ischinosiphon körn) fiber, which was recently found to present potential mechanical and ballistic properties as 30 vol% reinforcement of epoxy composites. To complement these properties, Izod impact tests are now communicated in this brief report for similar composites with up to 30 vol% of guaruman fibers. A substantial increase in impact resistance, with over than 20 times the absorbed energy for the 30 vol% guaruman fiber composite, was obtained in comparison to neat epoxy. These results were statistically validated by Weibull analysis, ANOVA, and Tukey’s test. Scanning electron microscopy analysis disclosed the mechanisms responsible for the impact performance of the guaruman fiber composites.

Molten globule state of human placental cystatin (HPC) at low pH conditions and the effects of trifluoroethanol (TFE) and methanol

2006 ◽  
Vol 84 (2) ◽  
pp. 126-134 ◽  
Author(s):  
Fouzia Rashid ◽  
Sandeep Sharma ◽  
M A Baig ◽  
Bilqees Bano
Keyword(s):  
Conformational Changes ◽  
Tertiary Structure ◽  
Molten Globule ◽  
Structural Features ◽  
Cd Spectroscopy ◽  
Native State ◽  
Molten Globule State ◽  
Fluorescence Measurements ◽  
Helical Content ◽  
Human Placental Cystatin

Acid-induced conformational changes were studied in human placental cystatin (HPC) in terms of circular dichroism (CD) spectroscopy, the binding of hydrophobic dye 1-anilinonapthalene-8-sulphonic acid (ANS), and intrinsic fluorescence measurements. Our results show the formation of an acid-induced molten globule state at pH 2.0, with significant secondary and tertiary interactions that resemble the native state, exposed hydrophobic regions and the effects of trifluoroethanol (TFE) and methanol in conversion of the acid-denatured state of HPC to the alcohol-induced state, which is characterized by increased helical content, disrupted tertiary structure, and the absence of hydrophobic clusters. Alcohol-induced formation of α-helical structures at pH 2.0 is evident from the increase in the ellipticity values at 222 nm, with native-like secondary structural features at 40% TFE. The increase in helical content was observed up to 80% TFE concentration. The ability of TFE (40%) to refold acid-denatured HPC to native-state conformation is also supported by intrinsic and ANS fluorescence measurements.Key words: human placental cystatin, molten globule, acid-induced state, trifluoroethanol, methanol, CD spectroscopy, ANS fluorescence, pH, protein folding.

Mimicking cotranslational folding of prosubtilisin E in vitro

2020 ◽  
Vol 167 (5) ◽  
pp. 473-482 ◽  
Author(s):  
Sung-Gun Kim ◽  
Yu-Jen Chen ◽  
Liliana Falzon ◽  
Jean Baum ◽  
Masayori Inouye
Keyword(s):  
Crucial Role ◽  
Tertiary Structure ◽  
Molten Globule ◽  
Secondary Structures ◽  
Terminal Region ◽  
Folding Intermediates ◽  
C Terminus ◽  
Cotranslational Folding ◽  
N Terminus

Abstract Nascent polypeptides are synthesized on ribosomes starting at the N-terminus and simultaneously begin to fold during translation. We constructed N-terminal fragments of prosubtilisin E containing an intramolecular chaperone (IMC) at N-terminus to mimic cotranslational folding intermediates of prosubtilisin. The IMC-fragments of prosubtilisin exhibited progressive enhancement of their secondary structures and thermostabilities with increasing polypeptide length. However, even the largest IMC-fragment with 72 residues truncated from the C-terminus behaved as a molten globule, indicating the requirement of the C-terminal region to have a stable tertiary structure. Furthermore, truncation of the IMC in the IMC-fragments resulted in aggregation, suggesting that the IMC plays a crucial role to prevent misfolding and aggregation of cotranslational folding intermediates during translation of prosubtilisin polypeptide.

Intermediate studies on refolding of arginine kinase denatured by guanidine hydrochloride

2005 ◽  
Vol 83 (2) ◽  
pp. 109-114 ◽  
Author(s):  
Hong-Min Tang ◽  
Hong Yu
Keyword(s):  
Fluorescence Intensity ◽  
Tertiary Structure ◽  
Structural Characteristics ◽  
Molten Globule ◽  
Guanidine Hydrochloride ◽  
Arginine Kinase ◽  
Fluorescence Emission ◽  
Tryptophan Fluorescence ◽  
Blue Shift ◽  
Molten Globule State

The refolding course and intermediate of guanidine hydrochloride (GuHCl)-denatured arginine kinase (AK) were studied in terms of enzymatic activity, intrinsic fluorescence, 1-anilino-8-naphthalenesulfonte (ANS) fluorescence, and far-UV circular dichroism (CD). During AK refolding, the fluorescence intensity increased with a significantly blue shift of the emission maximum. The molar ellipticity of CD increased to close to that of native AK, as compared with the fully unfolded AK. In the AK refolding process, 2 refolding intermediates were observed at the concentration ranges of 0.8–1.0 mol/L and 0.3–0.5 mol GuHCl/L. The peak position of the fluorescence emission and the secondary structure of these conformation states remained roughly unchanged. The tryptophan fluorescence intensity increased a little. However, the ANS fluorescence intensity significantly increased, as compared with both the native and the fully unfolded states. The first refolding intermediate at the range of 0.8–1.0 mol GuHCl/L concentration represented a typical "pre-molten globule state structure" with inactivity. The second one, at the range of 0.3–0.5 mol GuHCl/L concentration, shared many structural characteristics of native AK, including its secondary and tertiary structure, and regained its catalytic function, although its activity was lower than that of native AK. The present results suggest that during the refolding of GuHCl-denatured AK there are at least 2 refolding intermediates; as well, the results provide direct evidence for the hierarchical mechanism of protein folding.Key words: arginine kinase, guanidine-denatured, refolding, intermediate, molten globule state.

scholarly journals Synthesis and mechanical characterization of a non-woven nanofiber by the electrospinning technique

DYNA ◽  
2019 ◽  
Vol 86 (209) ◽  
pp. 64-72
Author(s):  
Edwin Edgardo Espinel Blanco ◽  
Nelson Escobar-Mora ◽  
Lina Marcela Hoyos-Palacio ◽  
Martin Fabián Sarmiento-Gaviria
Keyword(s):  
Electron Microscopy ◽  
Mechanical Characterization ◽  
Average Diameter ◽  
Electrospinning Technique ◽  
Acid Ratio ◽  
Nanostructured Polymer ◽  
Electron Microscopy Analysis ◽  
Microscopy Analysis ◽  
Scanning Electron

A non-woven nanofiber or polymeric cover is synthesized with nylon-6 as the base polymeric material. Different acid relationships (formic/acetic) were tested in the electrospinning equipment, until defining by macroscopic observations and SEM Scanning Electron Microscopy analysis the adequate acid ratio 3:2 and the average diameter of the nanofibers in 350nm, defining the parameters to operate the electrospinning. According to ASTM D 882 standard, the Tensile Strength was calculated for stresses applied horizontally and vertically to the direction of the nanofibers. With the standards ASTM D 7490 and ASTM G-15 the wettability was determined by measuring the contact angle, finding that it has hydrophilic properties with high wettability, adhesiveness and surface energy. Nanostructured polymer covers can be used for biological isolation in health care areas, as a protective barrier to control the spread of infections. 

Effect of Micro Scale Textures on Drilling Performance of Carbide Tools in Dry and Wet Machining of Ti-6Al-4V

2016 ◽  
Author(s):  
Niketh Saseendran ◽  
Samuel Gonumakulapalle Lodi
Keyword(s):  
Laser Scanning ◽  
Thrust Force ◽  
Average Diameter ◽  
Mechanical And Thermal Properties ◽  
Drill Bit ◽  
Drilling Performance ◽  
Electron Microscopy Analysis ◽  
Microscopy Analysis ◽  
Dry And Wet Conditions ◽  
Micro Dimples

Surface textures helps in controlling the tribological, optical, mechanical, and thermal properties on the surfaces. The recent advancements in precision machining, makes it possible to generate micro/nano patterns on the surfaces with high dimensional control. In the present work an attempt has been made to reduce the thrust force and torque developed during drilling super alloy, Ti-6Al-4V by creating micro dimples on the tool surfaces. Circular dimples having an average diameter of 35 micrometer were created on the flute and margin side of the drill bit using Nd:YAG laser. Scanning electron microscopy analysis has been done to evaluate the quality of generated micro dimples. Drilling experiments were carried out on the titanium alloy in both dry and wet conditions using flute textured, margin textured and non textured tool for understanding the effect of micro textures on the tool. From the force analysis it was observed that in both dry and wet conditions there was a considerable reduction in thrust force and torque. Surface inspections of the drill bit were performed using Stereomicroscope for investigating the titanium buildup on the cutting tool surfaces. Results showed that the margin textured tool performed better than the flute textured and untextured tool in both dry and wet conditions.

scholarly journals Molten Globule-Like Partially Folded State ofBacillus licheniformis α-Amylase at Low pH Induced by 1,1,1,3,3,3-Hexafluoroisopropanol

2014 ◽  
Vol 2014 ◽  
pp. 1-9 ◽  
Author(s):  
Adyani Azizah Abd Halim ◽  
Mohammed Suleiman Zaroog ◽  
Habsah Abdul Kadir ◽  
Saad Tayyab
Keyword(s):  
Thermal Denaturation ◽  
Tertiary Structure ◽  
Molten Globule ◽  
Low Ph ◽  
Intrinsic Fluorescence ◽  
Cd Spectra ◽  
Fluorescence Measurements ◽  
The Mean ◽  
Partially Folded State ◽  
Cooperative Transition

Effect of 1,1,1,3,3,3-hexafluoroisopropanol (HFIP) on acid-denaturedBacillus licheniformis α-amylase (BLA) at pH 2.0 was investigated by far-UV CD, intrinsic fluorescence, and ANS fluorescence measurements. Addition of increasing HFIP concentrations led to an increase in the mean residue ellipticity at 222 nm (MRE222 nm) up to 1.5 M HFIP concentration beyond which it sloped off. A small increase in the intrinsic fluorescence and a marked increase in the ANS fluorescence were also observed up to 0.4 M HFIP concentration, both of which decreased thereafter. Far- and near-UV CD spectra of the HFIP-induced state observed at 0.4 M HFIP showed significant retention of the secondary structures closer to native BLA but a disordered tertiary structure. Increase in the ANS fluorescence intensity was also observed with the HFIP-induced state, suggesting exposure of the hydrophobic clusters to the solvent. Furthermore, thermal denaturation of HFIP-induced state showed a non-cooperative transition. Taken together, all these results suggested that HFIP-induced state of BLA represented a molten globule-like state at pH 2.0.

Effect of composition and morphology of non-metallic inclusions on fracture toughness in as-cast AHSS

2019 ◽  
Vol 116 (6) ◽  
pp. 623
Author(s):  
Zhanfang Wu ◽  
Zhenyu Liu ◽  
Shengtao Qiu ◽  
Xiangyang Li
Keyword(s):  
Solidus Temperature ◽  
Calculation Model ◽  
Slow Cooling ◽  
Metallic Inclusions ◽  
Test Steel ◽  
Electron Microscopy Analysis ◽  
Chill Mold ◽  
Microscopy Analysis ◽  
Small Cracks ◽  
The Impact

A set of water-cooling copper-chill mold equipment is designed to study the precipitation behavior of different inclusion types in test steel under different cooling conditions, as well as its effects on the steel mechanical properties. As the results reveal, slow cooling treatment near the solidus temperature of test steel is conducive to forming more MnS + Al2O3 composite inclusions. The impact energy (−16 °C) of slow-cooled cast ingots at the core position is 28% higher than that of the air-cooled ingots. Scanning electron microscopy analysis of the specimens shows that small cracks are formed around single Al2O3 inclusions, but no cracks were found around composite inclusions. A calculation model of the tessellated stress of composite inclusions in steel is introduced to verify this phenomenon. The calculation demonstrates that, with a cover of ductile MnS, stress concentration around the composite inclusion is significantly decreased as compared with the single Al2O3 inclusion in the test steel.

SYNTHESIS OF MONODISPERSE Mn3O4 NANOCRYSTALS

2009 ◽  
Vol 08 (03) ◽  
pp. 281-283 ◽  
Author(s):  
MASOUD SALAVATI-NIASARI ◽  
FATEMEH DAVAR
Keyword(s):  
Photoelectron Spectroscopy ◽  
Average Diameter ◽  
Binding Energies ◽  
X Ray Diffraction ◽  
Transmission Electron Microscopy Analysis ◽  
X Ray ◽  
Transmission Electron ◽  
Electron Microscopy Analysis ◽  
Microscopy Analysis ◽  
20 Nm

Mn3O4 nanocrystals have been prepared using [bis(2-hydroxyacetophenato)manganese(II)] as precursor. Transmission electron microscopy analysis demonstrated nanocrystals Mn3O4 with an average diameter of about 20 nm. The structural study by X-ray diffraction indicates that these nanocrystals have pure tetragonal phase. The phase pure samples were characterized using X-ray Photoelectron Spectroscopy for Mn 2p level. The values of binding energies are consistent with the relative values reported in the literature.

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