DNA-dependent RNA polymerase in germinating bean rust uredospores

1973 ◽  
Vol 19 (9) ◽  
pp. 1175-1177
Author(s):  
M. S. Manocha
Keyword(s):  
Rna Polymerase ◽  
Ammonium Sulfate ◽  
Column Chromatography ◽  
Eukaryotic Cells ◽  
Bean Rust ◽  
Multiple Forms ◽  
Ammonium Sulfate Precipitation ◽  
Nucleoside Triphosphates ◽  
Dna Template

The multiple forms of DNA-dependent RNA polymerase normally obtained from eukaryotic cells are not found in the germinating uredospores of bean rust. Only one fraction peak of the enzyme is obtained after ammonium sulfate precipitation and column chromatography on DEAE-Sephadex (A-25). The enzyme of the germinating uredospores resembles RNA polymerase of procaryotic origin in its sensitivity to rifampicin, and in its requirements for a DNA template and for all four nucleoside triphosphates.

Radioimmunoassay for yeast killer toxin from Saccharomyces cerevisiae

1981 ◽  
Vol 27 (8) ◽  
pp. 847-849
Author(s):  
Farooq A. Siddiqui ◽  
Howard Bussey
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Ammonium Sulfate ◽  
Column Chromatography ◽  
Specific Activity ◽  
Killer Toxin ◽  
Ammonium Sulfate Precipitation ◽  
Antibody Preparation ◽  
Precipitation Band ◽  
Yeast Killer ◽  
Yeast Killer Toxin

Aradioimmunoassay was developed for the K1 killer toxin from strain T158C/S14a of Saccharomyces cerevisiae. 125I-labeled toxin was made to a specific activity of 100 μCi/mg of protein (1 μCi = 37 kBq). Antibody to purified toxin was prepared in rabbits using toxin cross-linked to itself. These antibodies, partially purified by 50% ammonium sulfate precipitation and Sepharose CL-6B column chromatography, produced one precipitation band with killer toxin and bound 125I-labeled toxin in a radioimmunoassay. The antibody preparation also bound with the toxins from another K1 killer, A364A, and three chromosomal superkiller mutants derived from it.

Partial characterization of the mode of inhibition of Escherichia coli RNA polymerase by the mixed disulfide, CoASSG

1979 ◽  
Vol 57 (4) ◽  
pp. 336-345 ◽  
Author(s):  
William C. H. Bees ◽  
Peter C. Loewen
Keyword(s):  
Escherichia Coli ◽  
Rna Polymerase ◽  
Equilibrium Dialysis ◽  
Mixed Disulfide ◽  
Nucleoside Triphosphates ◽  
Noncompetitive Inhibitor ◽  
Fe Complex ◽  
Short Time ◽  
Dna Template

The coenzyme A – glutathione mixed disulfide (CoASSG), when complexed with iron, is capable of inhibiting the RNA polymerase of Escherichia coli. A modified procedure involving a short time of exposure to high salt allowed the reliable preparation of CoASSG–Fe which was active in inhibiting RNA polymerase. The CoASSG–Fe complex acted as a noncompetitive inhibitor for the incorporation of all four nucleoside triphosphates but had a greater effect on GMP and CMP incorporation than AMP and UMP incorporation. Neither temperature nor ionic-strength changes affected CoASSG–Fe inhibition, and the use of rifampicin showed that CoASSG–Fe did not inhibit either the initiation or elongation processes of the polymerase. CoASSG–Fe was a more effective inhibitor at low DNA-template concentrations and it was more effective in inhibiting the incorporation of CMP and GMP on simple dG-dC containing templates and the asymmetric polymer poly d(T-C)∙poly d(G-A). The inhibition of transcription of poly d(I-C) was less effective than the inhibition of transcription of poly d(G-C). Equilibrium dialysis in microdialysis cells showed that CoASSG–Fe could associate with DNA in the absence of RNA polymerase.

Purification of Oidiodendron kalrai proteases

1976 ◽  
Vol 22 (3) ◽  
pp. 327-333 ◽  
Author(s):  
P. M. Cino ◽  
R. P. Tewari
Keyword(s):  
Ammonium Sulfate ◽  
Proteolytic Activity ◽  
Column Chromatography ◽  
Crude Extract ◽  
Proteolytic Enzymes ◽  
Deae Cellulose ◽  
Ammonium Sulfate Precipitation ◽  
Proteolytic Activities ◽  
Yeast Phase ◽  
Cellulose Column

Oidiodendron kalrai yeast-phase cells demonstrate proteolytic activity. Some of the proteolytic enzymes of the crude extract were purified by a combination of ammonium sulfate precipitation, Sephadex G-200, and diethylaminoethyl (DEAE) cellulose column chromatography. At least six proteins exhibiting a range of proteolytic activities could be identified by these procedures. Purity of the enzyme fractions obtained from the DEAE-cellulose columns was tested by running polyacrylamide gels.

Extraction and Partial Purification of Protease from Bilimbi (Averrhoa bilimbi L.)

2013 ◽  
Vol 10 (2) ◽  
pp. 29
Author(s):  
Normah Ismail ◽  
Nur' Ain Mohamad Kharoe
Keyword(s):  
Molecular Weight ◽  
Protein Content ◽  
Ammonium Sulfate ◽  
Molecular Weight Distribution ◽  
Weight Distribution ◽  
Protease Activity ◽  
Protein Concentration ◽  
Temperature Stability ◽  
Partial Purification ◽  
Ammonium Sulfate Precipitation

Unripe and ripe bilimbi (Averrhoa bilimbi L.) were ground and the extracted juices were partially purified by ammonium sulfate precipitation at the concentrations of 40 and 60% (w/v). The collected proteases were analysed for pH, temperature stability, storage stability, molecular weight distribution, protein concentration and protein content. Protein content of bilimbi fruit was 0.89 g. Protease activity of both the unripe and ripe fruit were optimum at pH 4 and 40°C when the juice were purified at 40 and 60% ammonium sulfate precipitation. A decreased in protease activity was observed during the seven days of storage at 4°C. Molecular weight distribution indicated that the proteases protein bands fall between IO to 220 kDa. Protein bands were observed at 25, 50 and 160 kDa in both the unripe and ripe bilimbi proteases purified with 40% ammonium sulfate, however, the bands were more intense in those from unripe bilimbi. No protein bands were seen in proteases purified with 60% ammonium sulfate. Protein concentration was higher for proteases extracted with 40% ammonium sulfate at both ripening stages. Thus, purification using 40% ammonium sulfate precipitation could be a successful method to partially purify proteases from bilimbi especially from the unripe stage. 

scholarly journals A set of simple methods for detection and extraction of laminarinase

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Ananthamurthy Koteshwara ◽  
Nancy V. Philip ◽  
Jesil Mathew Aranjani ◽  
Raghu Chandrashekhar Hariharapura ◽  
Subrahmanyam Volety Mallikarjuna
Keyword(s):  
Ammonium Sulfate ◽  
Ammonium Sulphate ◽  
Gold Standard ◽  
Direct Detection ◽  
Low Cost ◽  
Direct Analysis ◽  
Reducing Sugar ◽  
Ammonium Sulfate Precipitation ◽  
Simple Method ◽  
Functional Step

AbstractA carefully designed ammonium sulfate precipitation will simplify extraction of proteins and is considered to be a gold standard among various precipitation methods. Therefore, optimization of ammonium sulfate precipitation can be an important functional step in protein purification. The presence of high amounts of ammonium sulphate precludes direct detection of many enzymatically active proteins including reducing sugar assays (e.g. Nelson-Somogyi, Reissig and 3,5-dinitrosalicylic acid methods) for assessing carbohydrases (e.g. laminarinase (β (1–3)-glucanohydrolase), cellulases and chitinases). In this study, a simple method was developed using laminarin infused agarose plate for the direct analysis of the ammonium sulphate precipitates from Streptomyces rimosus AFM-1. The developed method is simple and convenient that can give accurate results even in presence of ammonium sulfate in the crude precipitates. Laminarin is a translucent substrate requiring the use of a stain to visualize the zones of hydrolysis in a plate assay. A very low-cost and locally available fluorescent optical fabric brightener Tinopal CBS-X has been used as a stain to detect the zones of hydrolysis. We also report simple methods to prepare colloidal chitin and cell free supernatant in this manuscript.

Preparation of Human Immunoglobulin by Ammonium Sulfate Precipitation

Vox Sanguinis ◽  
1976 ◽  
Vol 31 (6) ◽  
pp. 423-434
Author(s):  
A.F.S.A. Habeeb ◽  
Robert D. Francis
Keyword(s):  
Ammonium Sulfate ◽  
Human Immunoglobulin ◽  
Ammonium Sulfate Precipitation
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