An Endogenous Protease Activating Plasma Inactive Renin

1978 ◽  
Vol 55 (s4) ◽  
pp. 133s-134s ◽  
Author(s):  
B. J. Leckie
Keyword(s):  
Human Plasma ◽  
Serine Protease ◽  
Protease Inhibitors ◽  
Trypsin Inhibitor ◽  
Soya Bean ◽  
Acid Activation ◽  
Bean Trypsin Inhibitor ◽  
Inactive Renin ◽  
Inhibited Acid ◽  
Endogenous Protease

1. The protease inhibitors Trasylol and soya-bean trypsin inhibitor prevented the activation of plasma inactive renin by acid. 2. N-Ethylmaleimide inhibited acid-activation to some extent but o-phenanthroline had no effect. 3. Acid-activation of the inactive renin in human plasma is mediated by a serine protease.

Activation of latent collagenase from polymorphonuclear leukocytes by cathepsin G

1979 ◽  
Vol 44 (10) ◽  
pp. 3177-3182 ◽  
Author(s):  
Mária Stančíková ◽  
Karel Trnavský
Keyword(s):  
Affinity Chromatography ◽  
Trypsin Inhibitor ◽  
Polymorphonuclear Leukocytes ◽  
Soya Bean ◽  
Cathepsin G ◽  
Sepharose Column ◽  
Bean Trypsin Inhibitor ◽  
Human Polymorphonuclear Leukocytes

Cathepsin G was isolated from human polymorphonuclear leukocytes and purified by affinity chromatography on Antilysin-Sepharose column. Purified enzyme activated later collagenase isolated from leukocytes. Activation at 36°C was maximal after 30 min incubation. Inhibitors of cathepsin G - soya-bean trypsin inhibitor, diisopropyl phosphofluoridate and Antilysin were active in inhibiting the activation of latent collagenase by cathepsin G.

The Contribution of Plasma Protease Inhibitors to Antiplasmin Activity in Man

1976 ◽  
Vol 51 (2) ◽  
pp. 215-218
Author(s):  
G. P. M. Crawford ◽  
D. Ogston ◽  
A. S. Douglas
Keyword(s):  
Human Plasma ◽  
Protease Inhibitors ◽  
Trypsin Inhibitor ◽  
Antithrombin Iii ◽  
The Other ◽  
Plasmin Activity ◽  
Neutralizing Activity ◽  
Α2 Macroglobulin

1. Human plasma contains a variety of proteins that are capable of inhibiting plasmin activity. Whole plasma possesses ‘rapid’ and ‘progressive’ plasmin-neutralizing activity: this study assesses the contribution of individual protease inhibitors to this plasmin-neutralizing property of plasma. 2. Rapid and progressive antiplasmin activities of human plasma correlate with α2-macroglobulin and α1-antitrypsin concentrations respectively. 3. Fluctuations in the amounts of the other measured inhibitors (antithrombin III, Cl inactivator and inter-α-trypsin inhibitor) did not influence the measured antiplasmin activity.

The Nature of Inactive Renin in Human Plasma and Amniotic Fluid

1978 ◽  
Vol 55 (1) ◽  
pp. 41-50 ◽  
Author(s):  
A. A. Shulkes ◽  
R. R. Gibson ◽  
S. L. Skinner
Keyword(s):  
Molecular Weight ◽  
Amniotic Fluid ◽  
Human Plasma ◽  
Gel Filtration ◽  
Dilution Effect ◽  
Exchange Chromatography ◽  
Acid Activation ◽  
Temperature Dependent ◽  
Inactive Renin ◽  
Full Activation

1. The properties of inactive and active renin in human plasma and amniotic fluid were studied chromatographically. Activation was achieved at pH 3.3 with and without added pepsin. 2. Acid activation of renin was time- and temperature-dependent but was inhibited by dilution of the sample. The dilution effect was corrected by adding pepsin. Such characteristics indicate that activation at low pH is catalysed by intrinsic enzymes. 3. Separation and/or dilution of the activating enzyme during ion-exchange chromatography concealed the eluted position of inactive renin and reduced the amount recovered. Only after full activation of the eluted renin was achieved with added pepsin was a distinct peak of inactive renin exposed. 4. At pH 7.5 inactive renin carried a lower negative charge than the active enzyme. This charge difference was lost after activation. 5. No molecular-weight differences between active, inactive renin or the International Renin Standard were detected by gel filtration. No renin of larger molecular weight was present. 6. These findings will be helpful in purification studies of human inactive renin.

Effects of Soy Bean Trypsin Inhibitor on Fibrin Clot Lysis

1966 ◽  
Vol 15 (03/04) ◽  
pp. 542-553 ◽  
Author(s):  
K Egeblad
Keyword(s):  
Human Plasma ◽  
Blood Coagulation ◽  
Trypsin Inhibitor ◽  
Thrombin Generation ◽  
Inhibitory Effect ◽  
Fibrin Clot ◽  
Combined Effect ◽  
Clot Lysis ◽  
Bean Trypsin Inhibitor ◽  
Inhibitory Agents

SummaryThe effect of soy bean trypsin inhibitor (SBTI) on fibrinolysis and blood coagulation was investigated. Clot lysis was recorded by means of thrombelastography. SBTI delays fibrinolysis induced by plasmin and by activators of plasminogen (SK-activator and urokinase). Activator-induced lysis is delayed by a combined effect on activator and plasmin. There appears to exist an equilibrium between highly dissociated compounds of inhibitor with the active agents. The inhibitory effect of SBTI is relatively decreased in clots containing human plasma probably caused by an equilibrium with inhibitory agents in the plasma. SBTI delays thrombin generation in recalcified plasma as well as the effect of thromboplastin, but the effect is weak and requires a concentration 70-100 fold the antifibrinolytic active. The effect on thrombosis is even weaker.

scholarly journals Surface-simulation synthesis of the substrate-binding site of an enzyme. Demonstration with trypsin

1985 ◽  
Vol 226 (2) ◽  
pp. 477-485 ◽  
Author(s):  
M Z Atassi
Keyword(s):  
Binding Site ◽  
Trypsin Inhibitor ◽  
Binding Activity ◽  
Enzymic Activity ◽  
Soya Bean ◽  
Free Form ◽  
Protein Substrates ◽  
Substrate Analogues ◽  
Bean Trypsin Inhibitor ◽  
Alpha 1 Antitrypsin

From the X-ray co-ordinates of bovine trypsin and its complexes with substrate analogues (benzamidine) and with soya-bean trypsin inhibitor, a peptide (TP) was designed and synthesized by surface-simulation synthesis, a concept previously introduced by this laboratory, to mimic the binding site of trypsin. Also, a control peptide (CTP) was synthesized that contained all the amino acids present in the TP peptide, except that their order was randomized. The radioiodinated TP peptide bound specifically to adsorbents of benzamidine, whereas the control CTP peptide exhibited no binding activity. Conjugates to succinyl (3-carboxypropionyl)-lysozyme of the TP peptide, control CTP peptide and other unrelated peptides were examined by a radiometric binding assay for the ability to bind soya-bean trypsin inhibitor and human alpha 1-antitrypsin. Conjugates of the TP peptide exhibited considerable binding activity to adsorbents of soya-bean trypsin inhibitor or alpha 1-antitrypsin. None of the other peptide conjugates possessed any binding activity. Action of the active-site-directed reagents phenylmethanesulphonyl fluoride and di-isopropyl phosphorofluoridate on free TP and CTP peptides resulted in the modification of a serine residue in the TP peptide whereas the CTP peptide remained unaltered. The TP peptide, either in the free form or as a conjugate on succinyl-lysozyme, had no enzymic activity on protein substrates or on tosylarginine methyl ester. These findings indicated that the binding activity of an enzyme was well mimicked by the surface-stimulation peptide but that reproduction of the catalytic activity was not obtained.

scholarly journals Functional modifications of α2-macroglobulin by primary amines. Kinetics of inactivation of α2-macroglobulin by methylamine, and formation of anomalous complexes with trypsin

1982 ◽  
Vol 201 (1) ◽  
pp. 119-128 ◽  
Author(s):  
Fred Van Leuven ◽  
Jean-Jacques Cassiman ◽  
Herman Van Den Berghe
Keyword(s):  
Conformational Change ◽  
Trypsin Inhibitor ◽  
Binding Capacity ◽  
Soya Bean ◽  
Primary Amines ◽  
Slow Phase ◽  
Subunit Structure ◽  
Α2 Macroglobulin ◽  
Bean Trypsin Inhibitor ◽  
Kinetics Of

The unique steric inhibition of endopeptidases by human α2M (α2-macroglobulin) and the inactivation of the latter by methylamine were examined in relation to each other. Progressive binding of trypsin by α2M was closely correlated with the loss of the methylamine-reactive sites in α2M: for each trypsin molecule bound, two such sites were inactivated. The results further showed that, even at low proteinase/α2M ratios, no unaccounted loss of trypsin-binding capacity occurred. As α2M is bivalent for trypsin binding and no trypsin bound to electrophoretic slow-form α2M was observed, this indicates that the two sites must react (bind trypsin) in rapid succession. Reaction of [14C]methylamine with α2M was biphasic in time; in the initial rapid phase complex-formation with trypsin caused a largely increased incorporation of methylamine. In the subsequent slow phase trypsin had no such effect. These results prompted further studies on the kinetics of methylamine inactivation of α2M with time of methylamine treatment. It was found that conformational change of α2M and decrease in trypsin binding (activity resistant to soya-bean trypsin inhibitor) showed different kinetics. The latter decreased rapidly, following pseudo-first-order kinetics. Conformational change was much slower and followed complex kinetics. On the other hand, binding of 125I-labelled trypsin to α2M did follow the same kinetics as the conformational change. This discrepancy between total binding (125I radioactivity) and trypsin-inhibitor-resistant binding of trypsin indicated formation of anomalous complexes, in which trypsin could still be inhibited by soya-bean trypsin inhibitor. Further examination confirmed that these complexes were proteolytically active towards haemoglobin and bound 125I-labelled soya-bean trypsin inhibitor to the active site of trypsin. The inhibition by soya-bean trypsin inhibitor was slowed down as compared with reaction with free trypsin. The results are discussed in relation to the subunit structure of α2M and to the mechanism of formation of the complex.

Measurement of Endotoxin Levels in Plasma Using Limulus Amoebocyte Lysate and Chromogenic Substrate, S2222

1979 ◽  
Author(s):  
S Clark ◽  
M Scully ◽  
P Webb ◽  
V Kakkar
Keyword(s):  
Trypsin Inhibitor ◽  
Final Concentration ◽  
Soya Bean ◽  
Lag Phase ◽  
Chromogenic Substrate ◽  
Plasma Samples ◽  
Linear Calibration ◽  
Limulus Amoebocyte Lysate ◽  
Bean Trypsin Inhibitor ◽  
Pancreatic Trypsin Inhibitor

A method has been devised for the measurement of endotoxin in plasma using the chromogenic substrate, S2222, a substrate which has been shown to be particularly sensitive to the Limulus Lysate. Time curves of the rate of release of the chromogen in mixtures in which procoagulase activation was concurrent, were complex with a lag phase which was shortened by increasing endotoxin concentrations. At a final concentration of 0.5ng/ml and 370 activation was complete within 60 minutes. The enzyme was inhibited by soya bean trypsin inhibitor but not by pancreatic trypsin inhibitor or hirudin. In the method finally adopted the lysate (25µl) was incubated with endotoxin (E.coli 026.86 Difco) and magnesium chloride (final concentration 33mM) in a total volume of 225µl. After 12 minutes preincubation 165µl of S2222(0.4mM) was added and the increase in abdorbance at 405nm over two minutes measured using an Abbott Biochromatic Analyser 100. Linear assay curves were obtained with final concentration of 0.2 to 2.0ngs endotoxin/ml with ΔOD 405/min of 0.35 at 2.0ng endotoxin/ml of final incubation mixture. ΔOD /min in control tubes were of the order of 0.02. For measurement from plasma samples, endotoxin was first extracted with chloroform. Linear calibration curves were achieved at a concentration of endotoxin of 1 to 5ng/ml of whole blood with a net OD/min at the highest concentration of 0.25.

Luminal Epidermal Growth Factor is Trophic to the Small Intestine of Parenterally Fed Rats

1995 ◽  
Vol 89 (2) ◽  
pp. 117-120 ◽  
Author(s):  
Tania Marchbank ◽  
Robert A. Goodlad ◽  
Chung Y. Lee ◽  
Raymond J. Playford
Keyword(s):  
Small Intestine ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Parenteral Nutrition ◽  
Trypsin Inhibitor ◽  
Soya Bean ◽  
Trophic Effect ◽  
Bean Trypsin Inhibitor ◽  
Epidermal Growth ◽  
Intestinal Atrophy

1. Intestinal atrophy contributes to the clinical difficulties of patients on parenteral nutrition. Systemic administration of epidermal growth factor reverses this effect, but there is concern over the clinical safety of intravenous administration of growth factors. We therefore investigated whether administration of luminal epidermal growth factor could reverse the atrophy induced in a rat model of parenteral nutrition when epidermal growth factor was given alone or in combination with soya bean trypsin inhibitor to reduce proteolytic digestion of the epidermal growth factor. 2. Infusion of soya bean trypsin inhibitor alone decreased intraluminal tryptic activity by about 90% but did not result in increased proliferation. Intragastric infusion of epidermal growth factor (72 μg/day per rat) caused a 26% increase in proliferation (determined by 2-h metaphase arrest) in the duodenum (P < 0.01) when compared with animals receiving ‘control’ intragastric infusion. However, intragastric epidermal growth factor had no effect on more distal regions of the bowel, probably reflecting rapid proteolysis of the epidermal growth factor by luminal proteases. In contrast, a trophic effect of luminal epidermal growth factor was seen in the duodenum (28% increase, P < 0.01) and jejunum (24% increase, P < 0.05) of animals which had received epidermal growth factor with soya bean trypsin inhibitor. This was probably due to the soya bean trypsin inhibitor decreasing the rate of degradation of epidermal growth factor by intestinal proteases, allowing biologically active epidermal growth factor to reach more distal portions of the bowel. 3. We conclude that luminal administration of epidermal growth factor in combination with protease inhibitors could provide a novel approach to the treatment of intestinal atrophy induced by parenteral nutrition. This approach may also be useful for the treatment of ulceration of the small intestine in conditions such as necrotizing enterocolitis or Crohn's disease. Clinical studies are recommended.

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