scholarly journals CHARACTERIZATION OF VirB4 PROTEIN OF LOCAL ISOLATE Brucella abortus WITH WESTERN BLOTTING TECHNIQUE

2018 ◽  
Vol 12 (1) ◽  
Author(s):  
Ratih Novita Praja ◽  
Didik Handijatno ◽  
Setiawan Koesdarto ◽  
Aditya Yudhana
Keyword(s):  
Molecular Weight ◽  
Western Blotting ◽  
Virulence Factor ◽  
Brucella Abortus ◽  
Protein Band ◽  
Molecular Weights ◽  
Local Isolate ◽  
Blotting Technique

This research aimed to characterize VirB4 protein of local isolate Brucella abortus with Western blotting method. The result showed that there were four protein bands with molecular weights of 64.61, 59.25, 21.63, and 16.70 kDa by triggering a reaction between the whole Brucella abortus and anti-Brucella abortus serum. The results also revealed that there was only one protein band with a molecular weight of 59.25 kDa triggering a reaction between the whole Brucella abortus and anti-VirB Brucella abortus serum. Finally, it can be concluded that VirB4 protein can affect the virulence factor of Brucella abortus, successfully characterized with the appearance of one band with a molecular weight of 59.25 kDa by using Western blotting method.

scholarly journals Incorrect Molecular Weights due to inaccurate Prestained Protein Molecular Weight Markers that are used for Gel Electrophoresis and Western Blotting

2020 ◽  
Author(s):  
Rômulo Leão Silva Neris ◽  
Ajuni Kaur ◽  
Aldrin V. Gomes
Keyword(s):  
Molecular Weight ◽  
Molecular Mass ◽  
Western Blotting ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Protein Band ◽  
Molecular Weights ◽  
Molecular Masses ◽  
Protein Molecular Weight

ABSTRACTThe most widely used Western blotting protein standards are prestained proteins of known molecular mass (kDa). They are also utilized for sodium dodecyl sulphate (SDS) Polyacrylamide Gel Electrophoresis (PAGE) to determine the molecular mass of proteins separated by electrophoresis. The objective of this study was to assess the reliability of different commercially available protein standards in predicting accurate protein molecular weights. We performed this experiment by running Criterion TGX gels with five prestained protein standards (Thermo Fisher SeeBlue Plus 2, Bio-Rad Precision Plus Protein Dual-color, Thermo Fisher Spectra Multi-color, Novex-Sharp Pre-stained, and Invitrogen iBright Pre-Stained). To evaluate their accuracy, we utilized highly purified Bovine Serum Albumin (BSA, 66.44 kDa) and Cytochrome C (Cyto C, 11.62 kDa). We also made use of the dimers of BSA (132.88 kDa) and Cyt C (23.24 kDa) that are present on SDS-PAGE gels. Our results suggest that three of the standards were less accurate at higher molecular masses with the iBright marker having the highest error in determining the expected 132.88 kDa molecular weight. The SeeBlue Plus 2 was accurate at identifying the 132.88 kDa molecular weight protein band but was less reliable for the three other lower molecular weight proteins. These findings have significant implications for the determination of protein masses because researchers rely on these standards to evaluate the molecular masses of their protein(s). We suggest that at least two different protein standards should be initially used in electrophoresis gels and for Western blotting in order to get accurate protein molecular weight results.

scholarly journals α-Crystallin. The isolation and characterization of distinct macromolecular fractions

1971 ◽  
Vol 124 (2) ◽  
pp. 337-343 ◽  
Author(s):  
Abraham Spector ◽  
Lu-Ku Li ◽  
Robert C. Augusteyn ◽  
Arthur Schneider ◽  
Thomas Freund
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Chemical Difference ◽  
Molecular Weights ◽  
Isolation And Characterization ◽  
Different Populations ◽  
Molecular Weight Fractions

α-Crystallin was isolated from calf lens periphery by chromatography on DEAE-cellulose and gel filtration. Three distinct populations of macromolecules have been isolated with molecular weights in the ranges approx. 6×105−9×105, 0.9×106−4×106and greater than 10×106. The concentration of macromolecules at the molecular-weight limits of a population are very low. The members of the different populations do not appear to be in equilibrium with each other. Further, in those molecular-weight fractions investigated, no equilibrium between members of the same population was observed. The population of lowest molecular weight comprises 65–75% of the total material. The amino acid and subunit composition of the different-sized fractions appear very similar, if not identical. The only chemical difference observed between the fractions is the presence of significant amounts of sugar in the higher-molecular-weight fractions. Subunit molecular weights of approx. 19.5×103and 22.5×103were observed for all α-crystallin fractions.

scholarly journals Phaseolin type and heat treatment influence the biochemistry of protein digestion in the rat intestine

2008 ◽  
Vol 99 (3) ◽  
pp. 531-539 ◽  
Author(s):  
Carlos A. Montoya ◽  
Pascal Leterme ◽  
Stephen Beebe ◽  
Wolfgang B. Souffrant ◽  
Daniel Mollé ◽  
...  
Keyword(s):  
Heat Treatment ◽  
Western Blotting ◽  
Sole Source ◽  
Protein Band ◽  
Molecular Weights ◽  
Endogenous Protein ◽  
Anionic Trypsin ◽  
Endogenous Proteins ◽  
Protein Components

The study aimed to investigate thein vivodigestion ofPhaseolus vulgarisphaseolin types differing in their subunit pattern composition. Diets contained either casein as the sole source of protein or a mixture (1:1) of casein and pure Sanilac (S), Tendergreen (T) or Inca (I) phaseolin either unheated or heated. Rats were fed for 11 d with the experimental diets. Their ileal content and mucosa were collected and prepared for electrophoresis, Western blotting, densitometry and MS. Differences in digestion among native phaseolin types were observed for intact phaseolin at molecular weights (MW) of 47–50·5 kDa and for an undigested fragment at MW of 19–21·5 kDa in ileal digesta. In both cases, the concentration of these protein bands was lower for I phaseolin than for S or T phaseolin (P < 0·05). In the mucosa, the concentration of a protein band at MW of 20·5–21·5 kDa was lower for S phaseolin as compared to T or I phaseolin (P < 0·001). The presence of phaseolin subunits and their fragments was confirmed by Western blotting. MS analysis revealed the presence of undigested α and β subunit fragments from phaseolin and endogenous proteins (anionic trypsin I and pancreatic α-amylase) in ileal digesta. Thermal treatment improved digestion (P < 0·01), acting on both dietary and endogenous protein components. In conclusion, this study provides evidence for differences in intestinal digestion among phaseolin types, S phaseolin being more resistant and I phaseolin more susceptible. These differences were affected by the origin of the phaseolin subunit precursor. Heat treatment enhanced phaseolin digestion.

scholarly journals Isolation and Characterization of 2,3-Dichloro-1-Propanol-Degrading Rhizobia

2000 ◽  
Vol 66 (7) ◽  
pp. 2882-2887 ◽  
Author(s):  
Agus J. Effendi ◽  
Steven D. Greenaway ◽  
Brian N. Dancer
Keyword(s):  
Molecular Weight ◽  
High Ph ◽  
Molecular Weights ◽  
Isolation And Characterization ◽  
Complete Mineralization ◽  
Native Molecular Weight

ABSTRACT 2,3-Dichloro-1-propanol is more chemically stable than its isomer, 1,3-dichloro-2-propanol, and is therefore more difficult to degrade. The isolation of bacteria capable of complete mineralization of 2,3-dichloro-1-propanol was successful only from enrichments at high pH. The bacteria thus isolated were found to be members of the α division of the Proteobacteria in the Rhizobiumsubdivision, most likely Agrobacterium sp. They could utilize both dihaloalcohol substrates and 2-chloropropionic acid. The growth of these strains in the presence of 2,3-dichloro-1-propanol was strongly affected by the pH and buffer strength of the medium. Under certain conditions, a ladder of four active dehalogenase bands could be visualized from this strain in activity gels. The enzyme involved in the complete mineralization of 2,3-dichloro-1-propanol was shown to have a native molecular weight of 114,000 and consisted of four subunits of similar molecular weights.

Mechanical and Viscoelastic Characterization of Hyperbranched Polyisobutylenes

2002 ◽  
Vol 75 (5) ◽  
pp. 853-864 ◽  
Author(s):  
Judit E. Puskas ◽  
Christophe Paulo ◽  
Volker Altstädt
Keyword(s):  
Molecular Weight ◽  
Butyl Rubber ◽  
Structure Property ◽  
Carbocationic Polymerization ◽  
Molecular Weights ◽  
Structure Property Relationships ◽  
Molecular Weight Distributions ◽  
Weight Distributions ◽  
Living Carbocationic Polymerization

Abstract Structure-property relationships were investigated in hyperbranched polyisobutylenes, in comparison with commercial linear butyl rubber. The gel-free, soluble hyperbranched polyisobutylenes, synthesized by living carbocationic polymerization, had molecular weights, Mw≈400,000 to 1,000,000 g/mol, molecular weight distributions, MWD ≈1.2 to 2.6, and branching frequencies, BR ≈ 4 to 60. The mechanical and viscoelastic characterization of these polymers revealed interesting properties, including the characteristics of crosslinked rubbers.

Synthesis and Properties of Uniform Polyisoprene Networks. I. Synthesis and Characterization of α,ω-Dihydroxy-Polyisoprene

1976 ◽  
Vol 49 (2) ◽  
pp. 303-319 ◽  
Author(s):  
M. Morton ◽  
L. J. Fetters ◽  
J. Inomata ◽  
D. C. Rubio ◽  
R. N. Young
Keyword(s):  
Molecular Weight ◽  
Physical Properties ◽  
Soluble Fraction ◽  
Synthesis And Characterization ◽  
Molecular Weights ◽  
Molecular Weight Distributions ◽  
Weight Distributions ◽  
The Subject

Abstract The results of this study are the first to show that high-1,4 linear α,ω-dihydroxypolydienes can be synthesized with (a) predictable molecular weights, (b) narrow molecular weight distributions, and (c) high functionalities. Using the functionalized polyisoprenes prepared in this work, a series of networks was prepared with a purified triisocyanate as the chain linking agent. The soluble fraction in these networks ranged from 4.6 to 1.6 per cent. The characteristics and physical properties of these networks will be the subject of a forthcoming publication.

scholarly journals Characterization of proteins structurally related to human N-acetyl-β-d-glucosaminidase

1978 ◽  
Vol 173 (1) ◽  
pp. 191-196 ◽  
Author(s):  
M Carroll
Keyword(s):  
Molecular Weight ◽  
Human Liver ◽  
Deae Cellulose ◽  
Low Molecular Weight ◽  
Enzyme Preparations ◽  
Molecular Weights ◽  
Functional Relationships ◽  
Hexosaminidase A ◽  
Cellulose Column

Those proteins of human liver that cross-reacted with antibodies raised to apparently homogenous hexosamindases A and B were detected by immunodiffusion. Cross-reacting proteins with high molecular weights (greater than 2000000) and intermediate molecular weights (70000–200000) were present both in the unadsorbed fraction and in the 0.05–0.2M-NaCl eluate obtained by DEAE-cellulose chromatography at pH7.0. The unadsorbed fraction also contained a cross-reacting protein of low molecular weight (10000–70000). The possible structural and functional relationships between hexosaminidase and the cross-reacting proteins are discussed. An apparently cross-reacting protein present in the 0.05M-NaCl eluate from the DEAE-cellulose column was serologically unrelated to hexosaminidase, but it gave a reaction of immunological identify with one of the apparently cross-reacting proteins having the charge and size characteristics of hexosaminidase A. It is suggested that immunochemical methods may provide criteria for the homogeneity of enzyme preparations superior to those of conventional methods.

scholarly journals Direct Synthesis and Characterization of Photo-Crosslinkable Biodegradable PLA-PEG-PLA Ttiblock Copolymer with Methacrylates Functions by Green Montmorillonite Clay Catalyst

2020 ◽  
Vol 14 (4) ◽  
pp. 474-480
Author(s):  
Mohamed Benachour ◽  
◽  
Aslya El-Kebir ◽  
Amine Harrane ◽  
Rachid Meghabar ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Cationic Polymerization ◽  
Direct Synthesis ◽  
Average Molecular Weight ◽  
Montmorillonite Clay ◽  
Synthesis And Characterization ◽  
Molecular Weights ◽  
One Step ◽  
Clay Catalyst

Di-methacrylated PLA-PEG-PLA triblock copolymers of polylactide and polyethylene glycol were synthesized in one-step process by bulk cationic polymerization of lactide in the presence of PEG with different average molecular weights, using Maghnite-H+, an acidic montmorillonite clay, as a solid non-toxic catalyst. The obtained di-methacrylated copolymer was analyzed by 1H NMR and DSC. The effect of Maghnite-H+ proportions and PEG average molecular weight on the copolymerization and methacrylation yields and on average molecular weight of the resulting copolymers was studied.

scholarly journals ISOLATION AND PARTIAL CHARACTERIZATION OF MACRO- AND MICRONUCLEI FROM PARAMECIUM AURELIA

1972 ◽  
Vol 53 (1) ◽  
pp. 105-115 ◽  
Author(s):  
Donald J. Cummings
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Calcium Ions ◽  
Gel Electrophoresis ◽  
Partial Characterization ◽  
Intact Cells ◽  
Molecular Weights ◽  
High Molecular Weight Species ◽  
Nonionic Detergents

A method was developed for the isolation of macro- and micronuclei from Paramecium aurelia. This method utilized ionic and nonionic detergents to rupture the intact cells, calcium ions and spermidine were employed to protect the nuclei, and the nuclei were purified by centrifugation. Macronuclei consisted of 22% DNA, 10% RNA, and 68% protein. Micronuclei were composed of 9% DNA, 11% RNA, and 80% protein. DNA from both macro- and micronuclei had a density of 1.687 g/cc in CsCl and 1.417 g/cc in Cs2SO4. These values corresponded to G + C content of about 23%. The RNA of macronuclei was examined by gel electrophoresis, and two high molecular weight species were identified having molecular Weights of 1.3 x 106 and 2.8 x 106 daltons. Three syngens were studied, and in each case the conditions for isolation of the nuclei were the same and no differences were observed in the properties of the nuclei.

EXTRACTION AND CHARACTERIZATION OF HUMUS WITH REFERENCE TO CLAY-ASSOCIATED HUMUS

1974 ◽  
Vol 54 (3) ◽  
pp. 317-323 ◽  
Author(s):  
D. W. ANDERSON ◽  
E. A. PAUL ◽  
R. J. ST. ARNAUD
Keyword(s):  
Molecular Weight ◽  
Humic Acid ◽  
Gel Filtration ◽  
Spectral Measurements ◽  
Ultrasonic Dispersion ◽  
Agar Gel ◽  
Molecular Weights ◽  
Fractionation Method ◽  
C And N

An extraction-fractionation method was developed with which it is possible to isolate 60–67% of the humus of Chernozemic and Luvisolic soils. Two humic acid fractions were obtained: (1) a conventional alkali-pyrophosphate extractable HA-A; (2) a clay-associated HA-B fraction isolated after ultrasonic dispersion, in water, of the residue of the alkali-pyrophosphate extraction. As compared to the HA-A, the HA-B fractions had lower C contents but greater N contents, narrower C:H ratios, higher molecular weights, and less resistance to hydrolysis in 6 N HCl. It was concluded that the HA-B is a weakly humified, potentially labile humic constituent stabilized by adsorption to clay. Spectral measurements on low-ash (< 2%) humic acids showed a positive correlation between extinction coefficient at 280 nm (E280) and C:H ratio, and negative correlations between E280 and percent of hydrolyzable C and N. The E4:E6 ratio was related to molecular weight, as determined by agar gel filtration, increasing with decrease in molecular weight.

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