Isolation and partial characterization of bovine liver aminopeptidase B

1985 ◽  
Vol 50 (5) ◽  
pp. 1249-1257 ◽  
Author(s):  
Ján Blahovec ◽  
Michal Bartík ◽  
Evžen Kasafírek
Keyword(s):  
Molecular Weight ◽  
Gel Filtration ◽  
Bovine Liver ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Original Material ◽  
Purification Procedure ◽  
Michaelis Constant ◽  
Derivatives Of

Aminopeptidase B, specifically hydrolyzing the L-lysine and L-arginine derivatives of p-nitroaniline and β-naphthylamine, was isolated from bovine liver. A multistep purification procedure involving fractionation with ammonium sulfate, gel filtration on Sephadex, ion exchange chromatography on Ecteola-cellulose, and adsorption chromatography on hydroxylapatite, afforded an enzyme whose activity was approximately 240 times higher than the activity of the original material. The molecular weight of the enzyme determined by gel filtration on Sephadex G-200 was approximately 55 000. The Michaelis constant with respect to L-lysyl-p-nitroanilide was 1.2 . 10-3 mol/l.

Separation and Characterization of Two Fibrinolytic Inhibitors from Human Placenta

1971 ◽  
Vol 25 (03) ◽  
pp. 580-589 ◽  
Author(s):  
M Uszynski ◽  
U Abildgaard
Keyword(s):  
Molecular Weight ◽  
Ion Exchange ◽  
Isoelectric Focusing ◽  
Basic Protein ◽  
Gel Filtration ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Coagulation Inhibitor ◽  
Tissue Thromboplastin

SummaryProcedures for the separation of two inhibitors of the activation of plasminogen to plasmin by urokinase are described. Tissue thromboplastin was removed by adsorption to Al(0H)3 gel followed by ultracentrifugation. Plasminogen, plasminogen activator, a coagulation inhibitor and hemoglobin were removed by ion exchange chromatography (CM- or DEAE-Sephadex with NaCl gradients). The minor UK inhibitor is a relative basic protein with a pI of about 5.8. The major inhibitor was purified further by isoelectric focusing, preparative electrophoresis in polyacrylamide gel, and gel filtration. This inhibitor has α1-motility, the pI is about 5.2, and the molecular weight about 100,000. It inactivates urokinase progressively, but does not inhibit streptokinase, plasmin or thrombin.

Preparation and Characterization of NH2-Terminal Fibrinogen Bβ Fragments from N-DSK of Human Fibrinogen

1984 ◽  
Vol 51 (01) ◽  
pp. 016-021 ◽  
Author(s):  
S Birken ◽  
G Agosto ◽  
B Lahiri ◽  
R Canfield
Keyword(s):  
High Performance ◽  
Gel Filtration ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Reverse Phase ◽  
Human Fibrinogen ◽  
Human Volunteers ◽  
Cnbr Cleavage ◽  
Two Peaks

SummaryIn order to investigate the early release of NH2-terminal plasmic fragments from the Bβ chain of fibrinogen, substantial quantities of Bβ 1-42 and Bβ 1-21 are required as immunogens, as radioimmunoassay standards and for infusion into human volunteers to determine the half-lives of these peptides. Towards this end methods that employ selective proteolytic cleavage of these fragments from fibrinogen have been developed. Both the N-DSK fragment, produced by CNBr cleavage of fibrinogen, and Bβ 1-118 were employed as substrates for plasmin with the finding of higher yields from N-DSK. Bβ 1-42 and Bβ 1-21 were purified by gel filtration and ion-exchange chromatography on SP-Sephadex using volatile buffers. When the purified preparation of Bβ 1-42 was chromatographed on reverse-phase high performance liquid chromatography, two peaks of identical amino acid composition were separated, presumably due either to pyroglutamate or to amide differences.

scholarly journals The isocitrate dehydrogenases of Acinetobacter lwoffi. Separation and properties of two nicotinamide–adenine dinucleotide phosphate-linked isoenzymes

1972 ◽  
Vol 130 (1) ◽  
pp. 211-219 ◽  
Author(s):  
Colin H. Self ◽  
P. David J. Weitzman
Keyword(s):  
Molecular Weight ◽  
Ph Dependence ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Nicotinamide Adenine Dinucleotide Phosphate ◽  
Molecular Size ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Low Concentrations ◽  
Stimulation Of

Two isoenzymes of NADP-linked isocitrate dehydrogenase have been identified in Acinetobacter lwoffi and have been termed isoenzyme-I and isoenzyme-II. The isoenzymes may be separated by ion-exchange chromatography on DEAE-cellulose, by gel filtration on Sephadex G-200, or by zonal ultracentrifugation in a sucrose gradient. Low concentrations of glyoxylate or pyruvate effect considerable stimulation of the activity of isoenzyme-II. The isoenzymes also differ in pH-dependence of activity, kinetic parameters, stability to heat or urea and molecular size. Whereas isoenzyme-I resembles the NADP-linked isocitrate dehydrogenases from other organisms in having a molecular weight under 100000, isoenzyme-II is a much larger enzyme (molecular weight around 300000) resembling the NAD-linked isocitrate dehydrogenases of higher organisms.

Partial Primary Structure Of Human α2-Antiplasmin

1981 ◽  
Author(s):  
H R Lijnen ◽  
B Wiman ◽  
B Van Hoef ◽  
D Collen
Keyword(s):  
Amino Acids ◽  
Molecular Weight ◽  
High Performance Liquid Chromatography ◽  
High Performance ◽  
Gel Filtration ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Edman Degradation ◽  
Single Chain ◽  
Tryptic Digest

α2-Antiplasmin (α2AP), the main physiological inhibitor of plasmin in human plasma, is a single–chain glycoprotein with a molecular weight of 67,000 consisting of about 510 amino acids and containing 13 percent carbohydrate.A tryptic digest on 400 mg of reduced, carboxymethylated and citraconylated purified α2AP was performed. Peptides were separated by combinations of ion exchange chromatography, gel filtration and high performance liquid chromatography, and sequenced using the manual Edman degradation. Some peptides were further digested in order to establish overlaps. At the time of submission of this abstract we have sequenced 7 out of the approximately 21 arginyl peptides completely (each between 3 and 21 residues) and are working on the others. At present we have about 200 residues of sequence. Here we only report the stretches of 10 amino acids or more, which may be useful to compare the structure of α2AP with that of other serine protease inhibitors.

A dipeptidylpeptidase secreted by Fasciola hepatica

Parasitology ◽  
1994 ◽  
Vol 109 (1) ◽  
pp. 113-118 ◽  
Author(s):  
C. Carmona ◽  
S. McGonigle ◽  
A. J. Dowd ◽  
A. M. Smith ◽  
S. Coughlan ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Ion Exchange ◽  
Liver Fluke ◽  
Fasciola Hepatica ◽  
Gel Filtration ◽  
Serine Proteinase ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Substrate Preference ◽  
Proteolytic Digestion

SUMMARYA dipeptidylpeptidase (DPP) was isolated from Fasciola hepatica by gel-filtration and ion-exchange chromatography. The exoproteinase is secreted by newly excysted juveniles, immature and mature flukes. The liver fluke DPP is a serine proteinase of molecular weight > 200 kDa and differs from previously characterized mammalian DPPs in its substrate preference and susceptibility to inactivation by inhibitors. The parasite DPP may function in the latter stages of the proteolytic digestion of host macromolecules. In this manner, the enzyme may be important in providing the parasite with dipeptides that could be absorbed through the intestine as nutrient.

Enolase from Lobster (Homarus americanus)

1971 ◽  
Vol 28 (6) ◽  
pp. 879-882 ◽  
Author(s):  
M. John Chapman ◽  
Christopher Chin ◽  
Finn Wold
Keyword(s):  
Heat Treatment ◽  
Ion Exchange ◽  
Ammonium Sulfate ◽  
Catalytic Properties ◽  
Gel Filtration ◽  
Homarus Americanus ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Acetone Fractionation

Enolase has been isolated from lobster muscle by acetone fractionation, heat treatment, ammonium sulfate fractionation, gel filtration, and ion-exchange chromatography. Preliminary characterization of the pure enzyme shows that the catalytic properties are very similar to those of the enolases from rabbit and fish.

Purification and partial characterization of a plasma inhibitor of tonin

1981 ◽  
Vol 59 (4) ◽  
pp. 256-261 ◽  
Author(s):  
J. Tremblay ◽  
G. Thibault ◽  
J. Gutkowska ◽  
R. Boucher ◽  
J. Genest
Keyword(s):  
Molecular Weight ◽  
Plasma Proteins ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Sedimentation Velocity ◽  
Exchange Chromatography ◽  
Ammonium Sulfate Precipitation ◽  
Angiotensin I ◽  
Stokes Radius

A plasma inhibitor of tonin activity in the rat, was purified by ammonium sulfate precipitation, ion-exchange chromatography, and gel filtration. Its purity was investigated by analytical electrophoresis on polyacrylamide gel and by ultracentrifugation sedimentation velocity. The molecular weight (360 000) of the purified inhibitor was determined by sodium dodecyl sulfate electrophoresis and its isoelectric point (4.5) by gel isoelectrofocusing. The Stokes radius (640 nm) was evaluated by gel filtration studies and a frictional ratio (f/f0) of 1.95 was calculated from the molecular weight and Stokes radius.Kinetic studies using angiotensin I as substrate showed that the inhibition of tonin by the purified inhibitor was noncompetitive and does not exceed 70%. Electrophoresis showed the same mobility for [125I]tonin bound to plasma proteins and for [125I]tonin bound to the purified inhibitor. The inhibitor may be a protein resembling half of the dimeric protease inhibitor rat α1-macroglobulin or human α2-macroglobulin

scholarly journals High-Sulphur Proteins From a-Keratins ll. Isolation and Partial Characterization of Purified Components From Mouse Hair

1976 ◽  
Vol 29 (2) ◽  
pp. 11 ◽  
Author(s):  
Robert C Marshall ◽  
JM Gillespie
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Ion Exchange ◽  
Amino Acid Composition ◽  
Acid Composition ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Fractional Precipitation ◽  
Molecular Weight Range

The present paper continues the study of the reduced and S-carboxymethylated high-sulphur proteins from mouse hair. Fractions have been obtained in a substantially purified form by fractional precipitation with ammonium sulphate at pH 6, followed by ion exchange chromatography on cellulose phosphate at pH 2�6. Approximately 80% by weight of the high-sulphur proteins fall into the ultra-high-sulphur category (carboxymethyicysteine content greater than 26 residues per 100 residues), and they cover a molecular weight range of 17000-28000. The components show a remarkable diversity in amino acid composition; for example the contents of arginine and glycine each vary by about 3 : 1. The remainder of the proteins contain 17-20 residues per 100 residues of carboxymethyicysteine, are smaller in size (molecular weight 11 500), and also show great diversity in overall amino acid composition.

Purification and Properties of Human Factor IXa

1976 ◽  
Vol 35 (03) ◽  
pp. 576-585 ◽  
Author(s):  
Katalin Váradi ◽  
Susan Elödi
Keyword(s):  
Molecular Weight ◽  
Human Factor ◽  
Gel Filtration ◽  
Heat Stability ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Factor Ix ◽  
Clotting Factors ◽  
Factor Ixa ◽  
Purification And Properties

SummaryHuman factor IXa was purified 5,000-fold from serum by ion exchange chromatography. The preparation was free from other clotting factors. Both pH sensitivity and heat stability of purified factor IXa appeared to be different from those of factor IX in the plasma. The molecular weight of human factor IXa is 80,000 as estimated from gel-filtration experiments. Modification of seryl or histidyl side chains abolished the activity of factor IXa.

Characterisation of a Neutral Protease Produced by Micrococcus luteus

1996 ◽  
Vol 51 (5-6) ◽  
pp. 429-431 ◽  
Author(s):  
M.O. Ilori ◽  
O.O. Amund ◽  
O. Omidiji
Keyword(s):  
Molecular Weight ◽  
Citric Acid ◽  
Proteolytic Enzyme ◽  
Micrococcus Luteus ◽  
Gel Filtration ◽  
Apparent Molecular Weight ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Optimum Temperature ◽  
Optimum Ph

Abstract A proteolytic enzyme produced by a cassava-ferment­ing strain of Micrococcus luteus was extracted and puri­fied 50-fold by gel filtration and ion exchange chromatography. The optimum pH for the enzyme was 7.0, the optimum temperature 25 °C, the apparent molecular weight 42 kDa and the Km value, 0.45 mg ml-1 with casein as substrate. The enzyme was stimulated by Ca2+ and Mg2+ but inhibited by Zn2+ and Co2+ ions. Other inhibitors were EDTA, KCN, citric acid and L-cysteine indicating the enzyme to be a metalloprotease.

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