The self-association of glycosyl phenylazo dyes (Yariv antigens)

1978 ◽  
Vol 31 (10) ◽  
pp. 2225 ◽  
Author(s):  
EF Woods ◽  
GG Lilley ◽  
MA Jermyn
Keyword(s):  
Guanidine Hydrochloride ◽  
Sodium Dodecyl ◽  
Entropy Change ◽  
Wide Distribution ◽  
Enthalpy Change ◽  
Spectral Measurements ◽  
Concentrated Solutions ◽  
Weight Concentration ◽  
Equilibrium Sedimentation ◽  
Self Association

The aggregation of tris(4-glycosyloxyphenylazo)phloroglucinol compounds (Yariv antigens) where glyco = β-D-gluco, β-D-manno, β-D-galacto and α- D-galacto has been investigated by equilibrium sedimentation, the photoelectric scanning absorption optical system being utilized. The compounds have been shown to undergo a strong self-association in water and an analysis procedure employing Laplace transforms indicated a very wide distribution of species in the ultracentrifuge cell at equilibrium. To determine the type of association the concentration of monomer as a function of total weight concentration was calculated by using the recently developed Ω function. When the concentration of monomer is very low, as in the present case, the analysis is difficult because very slight curvature in the Ω-function plot has a large effect on the extrapolation to zero concentration. ��� The results are best described by an isodesmic indefinite self- association. The β-glucoside Yariv antigen was the most extensively investigated and the most highly purified sample was shown to have an association constant of 2.5 x 107 l. mol-1. The order of the association constants of the Yariv compounds was found to be β-glucoside > β- galactoside > α-galactoside > β-mannoside. For the β-glucoside compound an increase in temperature favours species of lower molecular weight, and from the enthalpy change a positive entropy change for the association is indicated. The compounds are disaggregated by concentrated solutions of urea and guanidine hydrochloride. Spectral measurements on the β-glucoside compound indicate disaggregation by sodium dodecyl sulfate above the critical micelle concentration.

scholarly journals Assessment of Conformational Changes in Low-Sulphur S-Carboxymethylkerateine from Wool

1967 ◽  
Vol 20 (4) ◽  
pp. 827 ◽  
Author(s):  
GM Bhatnagar ◽  
WG Crewther
Keyword(s):  
Absorption Spectrum ◽  
Conformational Changes ◽  
Guanidine Hydrochloride ◽  
Ultraviolet Absorption Spectrum ◽  
Spectral Measurements ◽  
Characteristic Difference ◽  
Concentrated Solutions ◽  
Difference Spectra ◽  
The Difference ◽  
Negative Difference

The effects of urea and guanidine hydrochloride on the ultraviolet absorption spectrum of the low-sulphur S-carboxymethylkerateine fraction of wool have been measured. In concentrated solutions of urea characteristic difference spectra were obtained with maxima of negative absorbance at 288, 280, and 240 miL. The addition of guanidine hydrochloride or an increase in temperature gave similar negative difference maxima at the higher wavelengths. Calculation of the extent of unfolding of the protein chains from the difference in absorbance at all three maxima showed that the unfolding was 50% complete at a urea concentration of about 1� 8M whereas a urea concep.tration of about 4� 3M was required to decrease the helix content by 50%. Similar measurements on components 7 and 8, the two major constituents of SCMKA, showed that a 50% decrease in helix content was obtained with 2�8M and O� 8M urea respectively whereas the corresponding values for 50 % unfolding assessed from difference spectral measurements were 2� 2M and 1� 2M urea respectively. It is suggested that the helical regions of components 7 and 8 aggregate specifically and that spectral measurements relate largely to non-helical portions of the chains.

scholarly journals A comparative study on the extraction effects of common agents on collagen-based binders in mural paintings

2021 ◽  
Vol 9 (1) ◽  
Author(s):  
Jianghao Du ◽  
Zhanyun Zhu ◽  
Junchang Yang ◽  
Jia Wang ◽  
Xiaotong Jiang
Keyword(s):  
Protein Structure ◽  
Comparative Study ◽  
Protein Extraction ◽  
Guanidine Hydrochloride ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Extraction Process ◽  
Ultraviolet Absorption Spectrum ◽  
Mural Paintings ◽  
The Impact

AbstractIn this paper, a comparative study was conducted on the extraction effects of six agents for collagen-based mural painting binders. These agents were used to extract the residual proteins in the non-aged and thermal aged samples. The protein extraction efficiencies of different extracting agents were quantitatively determined by bicinchoninic acid (BCA) method, and then processed by multivariate analysis of variance (MANOVA). The impact of the extraction process on the protein structure was characterized by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), ultraviolet absorption spectrum (UV) and circular dichroism (CD). The results showed that, for both non-aged and aged samples, the extraction efficiency of 2 M guanidine hydrochloride (GuHCl) was significantly higher than the other five agents, with less damage to the protein structure during the extraction process.

scholarly journals Macromolecular properties and polymeric structure of canine tracheal mucins

1991 ◽  
Vol 276 (2) ◽  
pp. 525-532 ◽  
Author(s):  
V Shankar ◽  
A K Virmani ◽  
B Naziruddin ◽  
G P Sachdev
Keyword(s):  
Gel Electrophoresis ◽  
Guanidine Hydrochloride ◽  
Polyacrylamide Gel Electrophoresis ◽  
Hydrophobic Properties ◽  
Experimental Conditions ◽  
Protein Core ◽  
Polymeric Structure ◽  
Composite Gel ◽  
Self Association ◽  
Static Laser Light Scattering

Two high-Mr mucus glycoproteins (mucins), CTM-A and CTM-B, were highly purified from canine tracheal pouch secretions, and their macromolecular properties as well as polymeric structure were investigated. On SDS/composite-gel electrophoresis, a diffuse band was observed for each mucin. Polyacrylamide-gel electrophoresis using 6% gels also showed the absence of low-Mr contaminants in the mucins. Comparison of chemical and amino acid compositions revealed significant differences between the two mucins. Using a static-laser-light-scattering technique, CTM-A and CTM-B were found to have weight-average Mr values of about 11.0 x 10(6) and 1.4 x 10(6) respectively. Both mucins showed concentration-dependent aggregation in buffer containing 6 M-guanidine hydrochloride. Under similar experimental conditions, reduced-alkylated CTM-A had an Mr of 5.48 x 10(6) and showed no concentration-dependent aggregation. Hydrophobic properties of the mucins, investigated by the fluorescent probe technique using mansylphenylalanine as the probe, showed the presence of a large number of low-affinity (KD approx. 10(5) M) binding sites. These sites appeared to be located on the non-glycosylated regions of the protein core, since Pronase digestion of the mucins almost completely eliminated probe binding. Reduction of disulphide bonds of CTM-A and CTM-B did not significantly alter the probe-binding properties. Also, addition of increasing NaCl concentrations (0.03-1.0 M) to the buffer caused only a small change in the hydrophobic properties of native and reduced-alkylated mucins. CTM-A was deglycosylated, without notable in the hydrophobic properties of native and reduced-alkylated mucins. CTM-A was deglycosylated, without notable degradation, using a combination of chemical and enzymic methods. On SDS/PAGE the protein core was estimated to have an Mr of approx. 60,000. On the basis of the protein and carbohydrate contents of the major mucin CTM-A, the mucin monomer was calculated to have an Mr of approx. 140,000. The high Mr (11 x 10(6] observed by physical methods is therefore due to self-association of the mucin monomer subunits.

Effect of detergents and chemicals on purified vaccinia virus: analysis by SDS polyacrylamide gel electrophoresis and electron microscopy

1977 ◽  
Vol 23 (3) ◽  
pp. 240-252 ◽  
Author(s):  
J. Boisvert ◽  
T. Yamamoto
Keyword(s):  
Electron Microscopy ◽  
Vaccinia Virus ◽  
Gel Electrophoresis ◽  
Alkali Treatment ◽  
Guanidine Hydrochloride ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Ammonium Bromide ◽  
Molecular Weights ◽  
Viral Particles

Vaccinia virus particles were dissociated into their constituent polypeptides and analysed by sodium dodecyl sulfate (SDS) gel electrophoresis. Thirty-three distinct polypeptide bands were identified and their molecular weights ranged between 11 000 and 150 000 daltons.Specific staining of gels containing polypeptides of dissociated virions revealed the presence of eight glycopeptides. No lipopeptides were detected.Analysis of chemical extracts (urea, guanidine hydrochloride, and alkali treatment) of the virus by SDS gel electrophoresis indicated that a total of 10 to 14 different polypeptides ranging in molecular weights from 11 000 to 70 000 daltons were solubilized.Analysis of detergent extracts and of the remains of extracted viral particles has shown that the detergent Nonidet P-40 (NP-40) solubilized a total of 11 polypeptides of which 6 were glycopeptides. The other detergents sodium deoxycholate (SDC) and cetyl trimethyl ammonium bromide (CTAB) were not as selective, both solubilizing more than 25 of the polypeptides composing the virus. Gel electrophoresis results also indicated that most of the small molecular weight (11 000–70 000 daltons) polypeptides were readily solubilized by NP-40, SDC, and CTAB, while those with molecular weights of 70 000 daltons and higher were not well solubilized.The effects of detergents were also analysed by electron microscopy. Evidence was obtained for subpopulations of viral particles having different susceptibility to detergent extraction.

scholarly journals Thermodynamics of ligand binding by the yeast mRNA-capping enzyme reveals different modes of binding

2004 ◽  
Vol 384 (2) ◽  
pp. 411-420 ◽  
Author(s):  
Isabelle BOUGIE ◽  
Amélie PARENT ◽  
Martin BISAILLON
Keyword(s):  
Ligand Binding ◽  
Entropy Change ◽  
Enthalpy Change ◽  
Mrna Capping ◽  
Capping Enzyme ◽  
Eukaryotic Mrnas ◽  
Entropic Effect ◽  
A Minor ◽  
Rna Capping ◽  
Capping Enzymes

RNA-capping enzymes are involved in the synthesis of the cap structure found at the 5′-end of eukaryotic mRNAs. The present study reports a detailed study on the thermodynamic parameters involved in the interaction of an RNA-capping enzyme with its ligands. Analysis of the interaction of the Saccharomyces cerevisiae RNA-capping enzyme (Ceg1) with GTP, RNA and manganese ions revealed significant differences between the binding forces that drive the interaction of the enzyme with its RNA and GTP substrates. Our thermodynamic analyses indicate that the initial association of GTP with the Ceg1 protein is driven by a favourable enthalpy change (ΔH=−80.9 kJ/mol), but is also clearly associated with an unfavourable entropy change (TΔS=−62.9 kJ/mol). However, the interaction between Ceg1 and RNA revealed a completely different mode of binding, where binding to RNA is clearly dominated by a favourable entropic effect (TΔS=20.5 kJ/mol), with a minor contribution from a favourable enthalpy change (ΔH=−5.3 kJ/mol). Fluorescence spectroscopy also allowed us to evaluate the initial binding of GTP to such an enzyme, thereby separating the GTP binding step from the concomitant metal-dependent hydrolysis of GTP that results in the formation of a covalent GMP–protein intermediate. In addition to the determination of the energetics of ligand binding, our study leads to a better understanding of the molecular basis of substrate recognition by RNA-capping enzymes.

The isolation of surface array proteins from bacteria

1984 ◽  
Vol 62 (11) ◽  
pp. 1181-1189 ◽  
Author(s):  
S. F. Koval ◽  
R. G. E. Murray
Keyword(s):  
Molecular Weight ◽  
Protein Conformation ◽  
Gel Electrophoresis ◽  
Guanidine Hydrochloride ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Structural Features ◽  
Proteolytic Degradation ◽  
Low Concentrations ◽  
Single Polypeptide

The methods used for the isolation of regularly structured (RS) surface array proteins of a range of prokaryotes are described. Most RS proteins can be selectively solubilized from envelope preparations with low concentrations of urea or guanidine hydrochloride. Sodium dodecyl sulfate – polyacrylamide gel electrophoresis analysis of the protein extracts shows that most RS arrays are composed of a single polypeptide that may contain carbohydrate. The molecular weight of the proteins varies from 41 000 to 200 000. Possible reasons for the presence of more than one polypeptide in RS protein preparations are discussed, as well as the evidence for proteolytic degradation of some RS proteins during isolation. Structural features of the RS proteins are described and the importance of protein conformation to assembly of the arrays is indicated.

scholarly journals A method for investigating the effect of temperature on the 695 nm band of insoluble cytochrome c

1975 ◽  
Vol 149 (1) ◽  
pp. 169-177 ◽  
Author(s):  
T A Moore ◽  
C Greenwood
Keyword(s):  
Ionic Strength ◽  
Cytochrome C ◽  
Computer Analysis ◽  
Standard Enthalpy ◽  
Entropy Change ◽  
Enthalpy Change ◽  
Effect Of Temperature ◽  
Standard Entropy ◽  
Ferricytochrome C ◽  
Standard Enthalpy Change

A method is described for computer analysis of simple spectrophotometric changes in particulate systems, and this has been applied to the bleaching of the 695 nm band of insoluble ferricytochrome c by temperature. The results show that insolubilization has no effect on the standard enthalpy change but lowers the value for the standard entropy change. This effect appears to be independent of the concentration of the gel matrix to which the cytochrome c is bound, but dependent on the ionic strength of the surrounding solution.

Isolation and preliminary characterization of histone H1.b allelic variants from quail erythrocytes

Genome ◽  
1998 ◽  
Vol 41 (5) ◽  
pp. 709-719 ◽  
Author(s):  
Jan Palyga ◽  
James M Neelin
Keyword(s):  
Guanidine Hydrochloride ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Cation Exchange Resin ◽  
Gel Permeation Chromatography ◽  
Histone H1 ◽  
Linker Histone ◽  
Allelic Variants ◽  
Limited Digestion

Our goal was to purify and characterize the allelic variants H1b1 and H1b2 of histone H1.b, one of the seven subtypes of this linker histone extracted from Japanese quail erythrocyte nuclei. These variants are revealed phenotypically as band H1.3 or part of band H1.4 by polyacrylamide gel electrophoresis (PAGE) in sodium dodecyl sulfate (SDS). All H1 subtypes together were separated from H5 by gel-permeation chromatography through Bio-Gel P-150. H1 was then fractionated on a column of the cation-exchange resin Amberlite CG-50 by using a shallow guanidine hydrochloride gradient, which enriched subtype H1.b together with H1.z and overlapping with subtypes H1.a and H1.b. Alternatively purification of subtypes was achieved electrophoretically: total H1 fractions from quail with different H1 phenotypes were first resolved into sub-types by PAGE in acetic acid - urea; after staining, the appropriate H1.b bands from several parallel gel pieces were excised and the histone was concentrated by PAGE in SDS. After fragmentation of H1.b in the gel pieces with N-bromosuccinimide (NBS), PAGE in SDS indicated no difference between H1b1 and H1b2 in the C-terminal "half" of the polypeptides. In contrast, limited digestion with endoprotease V8 from Staphylococcus aureus has shown that differences, probably by a few residues in length, reside in the N-terminal part of the molecule, close to the amino-terminus.Key words: quail histone H1.b, electrophoretic fractionation, linker histone variant.

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