scholarly journals The apparent molecular weights of human intestinal aminopeptidase, enterokinase and maltase in native duodenal fluid

1977 ◽  
Vol 165 (3) ◽  
pp. 583-585 ◽  
Author(s):  
A I Magee ◽  
D A W Grant ◽  
J Hermon-Taylor
Keyword(s):  
Molecular Form ◽  
Gel Chromatography ◽  
Duodenal Fluid ◽  
Molecular Weights ◽  
Induced Changes

The apparent molecular weights of human intestinal aminopeptidase, enterokinase and maltase in native duodenal fluid were estimated by gel chromatography on Sephadex G-200 under different conditions of operational buffer and temperature. No evidence for environmentally induced changes in molecular form was found.

Low Molecular Weight Trypsin-Plasmin Inhibitors Isolated from Papain Treated Urinary Trypsin Inhibitor

1982 ◽  
Vol 47 (01) ◽  
pp. 014-018 ◽  
Author(s):  
H Sumi ◽  
N Toki ◽  
S Takasugi ◽  
S Maehara ◽  
M Maruyama ◽  
...  
Keyword(s):  
Trypsin Inhibitor ◽  
Gel Electrophoresis ◽  
Specific Activity ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Chromatography ◽  
Urinary Trypsin Inhibitor ◽  
Plasmin Activity ◽  
Molecular Weights ◽  
Plasmin Inhibitors

SummaryPapain treatment of human urinary trypsin inhibitor (UTI67; mol. wt. 43,000 by SDS-polyacrylamide gel electrophoresis, specific activity 1,897 U/mg protein) produced four new protease inhibitors, which were highly purified by gel chromatography on Sephadex G-100 and isoelectric focusing. The purified inhibitors (UTI26, UTI9-I, UTI9-II, and UTI9-III) were shown to be homogeneous by polyacrylamide disc gel electrophoresis, and had apparent molecular weights of 26,000, 9,000, 9,000, and 9,800, respectively, by sodium dodecyl sulfate gel electrophoresis. During enzymatic degradation of UTI67, the amino acid compositions changed to more basic, and the isoelectric point increased from pH 2.0 (UTI67) to pHs 4.4, 5.2, 6.6, and 8.3 (UTI26, UTI9-I, UTI9-II, and UTI9-III), respectively. Both the parent and degraded inhibitors had anti-plasmin activity as well as antitrypsin and anti-chymotrypsin activities. Much higher anti-plasmin/anti-trypsin and anti-plasmin/anti-chymotrypsin activities were observed in the degraded inhibitors than in the parent UTI67. They competitively inhibited human plasmin with Ki values of 1.13 X 10-7 - 2.12 X 10-6 M (H-D-Val-Leu-Lys-pNA substrate). The reactions were very fast and the active site of the inhibitors to plasmin was thought to be different from that to trypsin or chymotrypsin.

scholarly journals Partial purification and properties of the two common inherited forms of human erythrocyte adenylate kinase

1972 ◽  
Vol 130 (3) ◽  
pp. 797-803 ◽  
Author(s):  
C. Brownson ◽  
N. Spencer
Keyword(s):  
Adenylate Kinase ◽  
Effect Of Temperature ◽  
Partial Purification ◽  
Heat Denaturation ◽  
Gel Chromatography ◽  
Molecular Weights ◽  
Purification And Properties ◽  
Increasing Temperature

1. The partial purification of adenylate kinase, types 1 and 2, from human erythrocytes is described. 2. Gel chromatography of both forms of the enzyme gave estimates of the molecular weights in the range 20000–23000. 3. Studies on crude haemolysates at various pH values indicated that the type 2 enzyme was less stable than the type 1. Heat denaturation studies on the partially purified enzymes confirmed these findings. 4. Measurements of rates of inhibition by iodoacetate and iodoacetamide showed that the type 2 enzyme reacts more readily than the type 1 enzyme with both reagents. 5. The effect of temperature on the initial velocity of ADP formation was measured at a single concentration of both AMP and MgATP2-. The two forms of the enzyme responded differently to increasing temperature.

Studies on Soluble Fibrin Complexes as a Function of pH

1975 ◽  
Author(s):  
Y. Benabid ◽  
E. Concord ◽  
M. Suscillon
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Gel Electrophoresis ◽  
Gel Chromatography ◽  
Agarose Gel ◽  
Monomer Mixture ◽  
Soluble Fibrin ◽  
Molecular Weights ◽  
Fibrin Monomer

Purified fibrinogen solutions, incubated with thrombin. CNBr. Sepharose, were subjected to agarose gel chromatography and eluted at different pH (6.5; 7.5; 8.5). Among high molecular weight derivatives formed by thrombin, the major component was a dimer. Gel chromatography at pH 8.5 showed a complexes peak distinct of that from fibrinogen, whereas at pH 6.5, only the fibrinogen peak appeared: fibrin monomer was eluted with fibrinogen as demonstrated by polyacrylamid gel electrophoresis 3.75% pH 8.9. SDS urea electrophoresis after reduction indicated that complexes peak contained two α-chains (α and α′). When fibrinogen was incubated with thrombin in the presence of FSF and calcium, several derivatives with higher and higher molecular weights were formed besides the dimer, and elution profiles of chromatography were identical at pH 6.5 and 8.5, thus indicating stable complexes formation. If fibrinogen-fibrin monomer mixture was subjected to FSF action at different pH, no complexes were formed at pH 6.5. These results confirm that at pH 6.5, any association was prevented.

Polysaccharides from the roots ofAlthaea officinalis L.: Structural features of D-glucans

1984 ◽  
Vol 49 (11) ◽  
pp. 2674-2679 ◽  
Author(s):  
Peter Capek ◽  
Rudolf Toman ◽  
Jozef Rosík ◽  
Alžbeta Kardošová ◽  
František Janeček
Keyword(s):  
Medicinal Plant ◽  
Structural Features ◽  
Gel Chromatography ◽  
Linear Character ◽  
Molecular Weights ◽  
Glycosidic Bonds ◽  
Althaea Officinalis ◽  
C Nmr

From the roots of the medicinal plant Althaea officinalis L., three D-glucans were isolated by gel chromatography which differed in molecular weights. The results of methylation analyses and 13C NMR measurements indicated predominantly linear character of the polysaccharide chains composed of α-D-glucopyranose units linked by 1 → 6 glycosidic bonds almost exclusively. The polymers had essentially the same structural features as D-glucan isolated from the leaves of this plant.

Isoprene and Rubber. Part 29. High Molecular Hydrorubbers

1932 ◽  
Vol 5 (2) ◽  
pp. 136-140
Author(s):  
H. Staudinger ◽  
W. Feisst
Keyword(s):  
Molecular Weight ◽  
Catalytic Reduction ◽  
Molecular Form ◽  
Average Molecular Weight ◽  
Molecular Size ◽  
Decomposition Products ◽  
Molecular Weights ◽  
Rubber Part ◽  
Derivatives Of ◽  
Suitable Process

Abstract The molecular concept in organic chemistry is based upon the fact that the molecules, whose existence is proved by vapor density determinations, enter into chemical reactions as the smallest particles. If now it is assumed that organic molecular colloids like rubber are dissolved in dilute solution in molecular form then it must be proved that in the chemical transposition of macromolecules as well no change in the size of the macromolecules occurs. In the case of hemicolloids, therefore for molecular colloids with an average molecular weight of 1000 to 10,000, this has been proved by the reduction of polyindenes, especially of polysterenes, to hydroproducts with the same average molecular weight, and also by the fact that cyclorubbers do not change their molecular weight upon autoöxidation. The molecular weights of hemi-colloidal hydrocarbons are therefore invariable. This is much more difficult to prove in the case of rubber, although there are many more ways in which unsaturated rubber can be transposed than the stable polysterenes, polyindenes, and poly cyclorubbers. In most of the reactions with rubber, as in its action with nitrosobenzene, oxidizing agents, hydrogen halides, and halogens, an extensive decomposition takes place as a result of the instability of the molecule, which is referred to in another work. Therefore derivatives of rubber are not formed, but derivatives of hemi-colloidal decomposition products. The catalytic reduction of rubber in the cold appears to be the most suitable process of making it react without changing its molecular size in order to prove that in a chemical transposition its molecular weight remains the same.

scholarly journals Cleavage of bacterial flagellin with cyanogen bromide. Chemical and physical properties of the protein fragments

1969 ◽  
Vol 113 (3) ◽  
pp. 489-499 ◽  
Author(s):  
C. R. Parish ◽  
G. L. Ada
Keyword(s):  
Amino Acid ◽  
Gel Electrophoresis ◽  
Amino Acid Analysis ◽  
Cyanogen Bromide ◽  
Degradation Products ◽  
Polyacrylamide Gel Electrophoresis ◽  
Polyacrylamide Gel ◽  
Gel Chromatography ◽  
Molecular Weights ◽  
Lysine Residues

1. Flagellin, isolated from the flagella of Salmonella adelaide, was shown by various criteria to be a pure protein. It had a molecular weight of about 40000 and contained three methionine, six tyrosine, 11 arginine and 25 lysine residues/mol., of which 11 of the lysine residues were present as ∈-N-methyl-lysine. 2. After treatment of flagellin with cyanogen bromide in formic acid, four main fragments (A, B, C and D) were obtained, with as many as six minor components that represented partial degradation products. The major fragments were estimated by amino acid analysis to have molecular weights of about 18000 for fragment A, 12000 for fragment B, 5500 for fragment C and 4500 for fragment D. Fragments A, B and D, but not fragment C, were recovered pure by gel chromatography as monitored by polyacrylamide-gel electrophoresis. 3. A complex between fragments C and D was also isolated (mol.wt. 10000) after limited oxidation of flagellin by chloramine-t before digestion by cyanogen bromide. After oxidation essentially only two fragments were released from flagellin by cyanogen bromide: the ‘C,D’ complex and a presumed ‘AB’ fragment. 4. The sum of the amino acid analyses of fragments A and B and the ‘C,D’ complex gave residue values that agreed well with the amino acid composition of native flagellin. 5. Fragments A and D contained tyrosine, and ten of the 11 ∈-N-methyl-lysine residues of the molecule were in fragment A. Reaction with [125I]iodide at small extents of substitution showed that, in flagellin, the tyrosine residue of fragment D was more readily substituted than those of fragment A. By contrast, in polymerized flagellin, the tyrosine residues of fragment A were more readily substituted. 6. Treatment of flagellin with carboxypeptidases A and B revealed the C-terminal sequence -Leu-Leu-Leu-Arg. Arginine and leucine were released by carboxypeptidase from the ‘C,D’ complex but not from fragment D, indicating that fragment C was C-terminal. 7. On the basis of the results from amino acid analysis, carboxypeptidase digestion, N-terminal analysis, iodination studies and polyacrylamide-gel electrophoresis, the sequence of fragments in flagellin was considered to be B–A–D–C; in the polymer, fragment A was exposed. It is suggested that methylation of the lysine residues occurred in the organism after flagellin had polymerized.

scholarly journals Plasmin Inhibitors in Human Urine

1977 ◽  
Author(s):  
H. Sumi ◽  
Y. Takada ◽  
A. Takada
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Human Urine ◽  
Molecular Form ◽  
Membrane Filter ◽  
Gel Filtration ◽  
Total Activity ◽  
Native Form ◽  
Molecular Weights ◽  
Two Peaks

By the gel filtration of urinary protein on Sephadex G-200, two peaks of activity to hydrolyze Nα-acetylglycyl-L-lysine methyl ester (AGLMe) were detected. One was the native form of urokinase, and the molecular weight was about 54,000. The other was of high molecular weight, and eluted in void fraction. The high molecular form was thought to be a complex of urokinase and urinary plasmin inhibitor (UPI). By using Arg-Sepharose, membrane filter (M.W. 10,000), and pevikon block electrophoresis, we could isolate four types of UPI from normal human urine. One UPI was positively charged at pH 8.6, and of high molecular weight. Other types were negatively charged, and the molecular weights by gel filtration on Sephadex G-200 were about 67,000 (UPI6.7), 45,000 (UPI4.5), and 22, 000 (UPI2.2), respectively. In acrylamide disc gel electrophoresis, UPI57 migrated to serum prealbumin fraction, and UPI45 and UPI? 2 were less anionic. Negative-charged UPIs could be adsorbed on trypsin-Sepharose, and were thought to be identical to urinary trypsin inhibitors. Purified UPIs showed strong inhibition on caseinolytic- and esterolytic-activities of plasmin, and the total activity was about 16 UPIU(inhibited 16 casein U of plasmin)/liter of urine.

scholarly journals Purification and Partial Characterization of Rat Fibrinogen (rat-Fbg)

1977 ◽  
Author(s):  
W. Nieuwenhuizen ◽  
Irina A.M. van Ruijven-Vermeer
Keyword(s):  
Amino Acid ◽  
Gel Electrophoresis ◽  
Proteolytic Enzymes ◽  
Mammalian Species ◽  
Rat Plasma ◽  
Gel Chromatography ◽  
Molecular Weights ◽  
Ammonium Sulphate Precipitation ◽  
Proteolytic Activities

Rat-Fbg was purified from rat plasma using Sepharose-1ysi ne chromatography, repeated ammonium sulphate precipitation (35% saturation) and gel chromatography on Sepharose 6B.In order to minimize proteolytic activity, rats were injected intravenously with Trasylol before bleeding, and the collected blood was treated with Trasylol and DFP.A preparation was obtained, which was 95% clottable and showed a single band on SDS-poly-acrylamide gel electrophoresis. Alanine was the only detectable am i no-term i na 1 amino acid.After reduction and modification of the SH groups the material could be separated into three distinct chains (Aα, Bβ and γ) by pore-limit Polyacrylamide slab-gel electrophoresis in sod i urn dodecyl sulphate. The amino acid composition of the whole Fbg and of the separated modified chains were determined. The molecular weights were 61,000, 58,000 and 51,000 for Aα, Bβ and γ-chains, respectively.In as far as the chains are concerned, our results are in contrast with the findings of Bouma et al. (J. Biol. Chem. 250(1975) 4678), who could not discriminate between Aα- and Bβ-chains in SDS-polyacry1 am i de gel electrophoresis. Evidence will be presented that this can be due to Aa-chain degradation caused by incomplete inhibition of proteolytic enzymes during the purification.It is concluded, that complete inhibition of proteolytic activities in all purification steps is essential to obtain native fibrinogen. Moreover, in contrast to the conclusions of Bouma rat-Fbg does not differ essentially from Fbg from other mammalian species.

THE EFFECTS OF THE PAI DERIVED FROM HUMAN PLATELETS ON TWO DIFFERENT TYPES OF t-PA

1987 ◽  
Author(s):  
T Yasukouchi ◽  
T Fujie ◽  
S Sakurama ◽  
M Satoh ◽  
M Ikeo ◽  
...  
Keyword(s):  
Molecular Form ◽  
Platelet Rich Plasma ◽  
Human Platelets ◽  
Red Cross ◽  
Molecular Weights ◽  
Rich Solution ◽  
The Difference ◽  
Triton X ◽  
Important Significance ◽  
Short Incubation Time

We studied on the effect of PAI derived from human platelets on two different kinds of t-PAs; one was purchased from BioPool, Sweden, (human uterus or melanoma cell derived one-chain t-PA: one-chain u-mt-PA) and another was kindly supplied from Sumitomo Pharm. Co., Osaka, (recombinant two-chain t-PA: two-chain rt-PA). As the PAI-rich solution derived from platelets. We used the platelets extract, which was prepared as follows: The concentrated platelet-rich plasma (Red Cross Japan) was washed with phosphate buffer containing 0.4 % Triton X-100. The inhibitory activities of the PAI were measured by the method of parabolic rate assay and were calcurated from the activities remained in the mixtures of PAI and the t-PAs. The reactions of the PAI and the t-PAs were also analysed by the method of fibrin autography. The PAI suppressed the activity of two-chain rt-PA completely within 5 minutes, but could not suppressed that of one-chain u-mt-PA completely within such a short incubation time. These facts also recognized on fibrin autography. The molecular weights of both free t-PAs were recognized at 60 kDa and that of the complex of the PAI and two-chain rt-PA was recognized at 110 kDa, respectively. The fibrinolytic zone of the complex of PAI and one-chain u-mt-PA, however, could not be recognized. This indicates two possibilities; one is too small amounts of complex forming to detect and another is that SDS can not reveal the t-PA activity from the complex of PAI and one-chain u-mt-PA on fibrin autography. It remained unclear that the difference of the reactions of these t-PAs to PAI had been induced from whether the difference between one- and two-chain, or that between rt-PA and u-mt-PA. If the former is reasonable, it can be said that the molecular form of t-PA has an important significance in physiological fibrinolytic mechanism.

Sign in / Sign up

Export Citation Format

Share Document