scholarly journals A simple procedure for the isolation of protein disulphide-isomerase

1987 ◽  
Vol 247 (1) ◽  
pp. 237-239 ◽  
Author(s):  
J Koivu ◽  
R Myllylä ◽  
K I Kivirikko
Keyword(s):  
Ion Exchange ◽  
Affinity Chromatography ◽  
Simple Procedure ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Beta Subunit ◽  
Protein Disulphide Isomerase ◽  
Step Procedure ◽  
Beta 2

A two-step procedure is described for the purification of protein disulphide-isomerase (PDI). This procedure is based on the previous finding that the beta-subunit of the prolyl 4-hydroxylase tetramer (alpha 2 beta 2) is identical with PDI [Koivu, Myllylä, Helaakoski, Pihlajaniemi, Tasanen & Kivirikko (1987) J. Biol. Chem. 262, 6447-6449; Pihlajaniemi, Helaakoski, Tasanen, Myllylä, Huhtala, Koivu & Kivirikko (1987) EMBO J. 6, 643-649]. The procedure involves purification of the prolyl 4-hydroxylase tetramer by a simple affinity chromatography and subsequent isolation of the beta-subunit from the dissociated tetramer by ion-exchange chromatography.

scholarly journals Human β-adrenergic receptors. Simultaneous purification of β1- and β2-adrenergic-receptor peptides

1987 ◽  
Vol 248 (2) ◽  
pp. 557-566 ◽  
Author(s):  
S W Bahouth ◽  
C C Malbon
Keyword(s):  
Ion Exchange ◽  
Affinity Chromatography ◽  
Adrenergic Receptors ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Beta Adrenergic Receptors ◽  
Beta Adrenergic ◽  
Hydrophobic Chromatography ◽  
Steric Exclusion ◽  
Beta 2

Beta-Adrenergic receptors from basal membranes of human placenta were purified from digitonin extracts by sequential rounds of affinity chromatography, hydrophobic chromatography, ion-exchange chromatography and steric-exclusion h.p.l.c. Basal membranes display both beta 1- and beta 2-adrenergic receptors, in the ratio 65:35. Affinity chromatography, hydrophobic chromatography on heptylamine-Sepharose and ion-exchange chromatography on DEAE-Sephacel removed most of the contaminating proteins, and final purification of the receptor to apparent homogeneity was achieved by steric-exclusion h.p.l.c. The purified receptors showed Mr 67000 on SDS/polyacrylamide-gel electrophoresis under reducing conditions. Specific binding of radioligand to the purified beta-adrenergic receptors displayed stereoselectivity, and the agonist competition profiles demonstrated the presence of both beta 1- and beta 2-receptors. By using the subtype-selective ligands CGP-20712A (beta 1-selective) and ICI-118,551 (beta 2-selective), the purified Mr-67000 species was shown to be composed of equivalent amounts of beta 1- and beta 2-adrenergic receptors. Affinity chromatography on Sepharose-alprenolol and sequential elution with 1 microM-CGP-20712A followed by 100 microM(-)-alprenolol permitted beta 1-adrenergic receptors to be resolved from the mixture of beta 1-/beta 2-adrenergic receptors. The pharmacologically distinct human beta 1 and beta 2-adrenergic receptors are shown to be structurally very similar peptides.

scholarly journals Comparison of the Affinity Chromatography and the Ion Exchange Chromatography in the Isolation of Bovine Immunoglobin G

OALib ◽  
2014 ◽  
Vol 01 (06) ◽  
pp. 1-5
Author(s):  
Mrigendra Rajput ◽  
Shimaa M. G. Mansour ◽  
Lyle J. Braun ◽  
Mahmoud Darweesh ◽  
Neelu Thakur ◽  
...  
Keyword(s):  
Ion Exchange ◽  
Affinity Chromatography ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Immunoglobin G

LINGCOD MUSCLE PHOSPHORIBOISOMERASE AND RIBULOSE 5′-PHOSPHATE 3′-EPIMERASE

1959 ◽  
Vol 37 (8) ◽  
pp. 961-973 ◽  
Author(s):  
H. L. A. Tarr
Keyword(s):  
Acid Phosphatase ◽  
Ion Exchange ◽  
Ammonium Sulphate ◽  
Simple Procedure ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Water Extraction ◽  
Enzyme Preparations

Comparatively pure phosphoriboisomerase and ribulose 5′-phosphate 3′-epimerase enzyme preparations were obtained from lingcod muscle by a simple procedure involving water extraction, saturation of the extract with ammonium sulphate, dialysis, brief heating to 55 °C, lyophilization of the solution, and final separation by ion exchange chromatography, using diethylamiuoethyl cellulose columns. Both enzymes have broad pH optima above pH 7.0, but are rapidly inactivated below this value. The following equilibria were established and compared with those obtained by other investigators: [Formula: see text], 1.35:1.0; [Formula: see text], 1: 1.5 and [Formula: see text], 1:0.58:0.66. The ketopentulose phosphate resulting from the action of phosphoriboisomerase on D-ribose 5-phosphate was isolated and identified as D-ribulose5-phosphate. Both D-ribulose and D-xylulose were demonstrated after subjecting a product of epimerase action to hydrolysis by acid phosphatase and ion exchange chromatography.

scholarly journals Current Trends in Protein Purification : A Review

2020 ◽  
pp. 279-310
Author(s):  
Angela Boxi ◽  
Isha Parikh ◽  
Radhika B S ◽  
Shryli K S
Keyword(s):  
High Pressure ◽  
Liquid Chromatography ◽  
Ion Exchange ◽  
Affinity Chromatography ◽  
Protein Purification ◽  
Gel Filtration ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Current Trends ◽  
Comparative Information

The present review is based on papers published between 1990 and 2020 and gives Comparative information about the most common protein purification techniques Gel-Filtration, Chromatography, Ion-Exchange Chromatography, Electrophoresis, Affinity Chromatography, and Dialysis, High-Pressure Liquid Chromatography. and their applications.

scholarly journals Thiol-protein disulphide oxidoreductases. Differences between protein disulphide-isomerase and glutathione-insulin transhydrogenase activities in ox liver

1976 ◽  
Vol 159 (2) ◽  
pp. 385-393 ◽  
Author(s):  
H C Hawkins ◽  
R B Freedman
Keyword(s):  
High Resolution ◽  
Ion Exchange ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Heat Denaturation ◽  
Differential Centrifugation ◽  
Protein Disulphide Isomerase ◽  
Final Sample ◽  
Thiol Protein ◽  
Single Enzyme

1. Protein disulphide-isomerase and glutathione-insulin transhydrogenase activities were assayed in parallel through a conventional purification of protein disulphide-isomerase from ox liver. 2. Throughout a series of purification steps (differential centrifugation, acetone extraction, (NH4)2SO4 precipitation and ion-exchange chromatography), the two activities appeared in the same fractions but were purified to different extents. 3. The final sample was 143-fold purified in protein disulphide-isomerase but only 10-fold purified in glutathione-insulin transhydrogenase; nevertheless the two activities in this preparation were not resolved by high-resolution isoelectric focusing and both showed pI4.65. 4. In a partially purified preparation containing both activities, glutathione-insulin transhydrogenase was far more sensitive to heat denaturation than was protein disulphide-isomerase; conversely protein disulphide-isomerase was more sensitive to inactivation by deoxycholate. 5. The data are inconsistent with a single enzyme being responsible for all the protein disulphide-isomerase and glutathione-insulin transhydrogenase activity of ox liver. It is suggested that several similiar thiol-protein disulphide oxidoreductases of overlapping specificities may better account for the data.

scholarly journals Bovine liver thiol-protein disulphide oxidoreductases. An alternative method for differential purification and resolution of protein disulphide-isomerase and glutathione-insulin transhydrogenase

1980 ◽  
Vol 191 (2) ◽  
pp. 389-393 ◽  
Author(s):  
D A Hillson ◽  
R B Freedman
Keyword(s):  
Ion Exchange ◽  
Rapid Method ◽  
Bovine Liver ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Protein Disulphide Isomerase ◽  
The Relationship ◽  
Thiol Protein ◽  
Homogeneous Preparation ◽  
Covalent Chromatography

1. Protein disulphide-isomerase (EC 5.3.4.1) and glutathione-insulin transhydrogenase (EC 1.8.4.2) activities in bovine liver were studied in parallel during purification of ‘thiol-protein disulphide oxidoreductase’ by the procedure of Carmichael, Morin & Dixon [(1977) J Biol. Chem. 252, 7163-7167]. The two activities showed no quantitative co-purification and were partially resolved by (NH4)SO4 precipitation, indicating that distinct enzymes are present. 2. Protein disulphide-isomerase was purified by a relatively rapid method involving a combination of the early stages of the Carmichael procedure and covalent chromatography, with a new stepwise elution procedure. Ion-exchange chromatography yields a homogeneous preparation of mol.wt. 57 000. 3. The relationship between protein disulphide-isomerase, glutathione-insulin transhydrogenase and ‘thiol-protein disulphide oxidoreductase’ is discussed.

Effects of whole blood storage on results for glycosylated hemoglobin as measured by ion-exchange chromatography, affinity chromatography, and colorimetry.

1983 ◽  
Vol 29 (6) ◽  
pp. 1113-1115 ◽  
Author(s):  
R R Little ◽  
J D England ◽  
H M Wiedmeyer ◽  
D E Goldstein
Keyword(s):  
Ion Exchange ◽  
Affinity Chromatography ◽  
Whole Blood ◽  
High Performance ◽  
Glycosylated Hemoglobin ◽  
Thiobarbituric Acid ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Sample Collection ◽  
Sample Stability

Abstract After storage of whole blood at either 4 or 20 degrees C, results for glycosylated hemoglobin by ion-exchange chromatography ("high-performance" liquid and mini-column chromatography), thiobarbituric acid colorimetry, and affinity chromatography were compared. At 4 degrees C, all methods gave acceptable results for samples stored for as long as a week. At 20 degrees C, the colorimetric and affinity methods also showed sample stability for a week or more. The ion-exchange methods were associated with a marked increase in values for glycosylated hemoglobin after a few days of storage. Evidently, care in details of sample collection and handling is especially important for ion-exchange methods, and the colorimetric and affinity methods have advantages over ion exchange in situations where long delays between sample collection and assay are unavoidable.

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