New fluorogenic substrates for the determination of post-proline endopeptidase activity

1990 ◽  
Vol 55 (4) ◽  
pp. 1112-1118 ◽  
Author(s):  
Chryssa Tzougraki ◽  
George Kokotos ◽  
Hana P. Mašková ◽  
Eva Anzenbacherová ◽  
Tomislav Barth
Keyword(s):  
Enzyme Activity ◽  
Organic Solvent ◽  
Fluorogenic Substrate ◽  
Michaelis Constant ◽  
Fluorogenic Substrates ◽  
Endopeptidase Activity ◽  
Optimum Ph ◽  
Adverse Influence

A new fluorogenic substrate for determination of the activity of post-proline endopeptidase (EC 3.4.21.26), Z-Cys(Bzl)-Pro-NH-Meq has been synthesized. Affinity of this substrate to the enzyme was significantly higher than that of the hitherto employed substrates. The Michaelis constant of the post-proline endopeptidase for Z-Cys(Bzl)-Pro-NH-Meq was at the optimum pH (7.0) 1.05 10-6 mol l-1. The concentration of dimethylsulfoxide used for the solubilization of Z-Cys(Bzl)-Pro-NH-Meq (<0.4%) was outside the limits of adverse influence of the organic solvent on the enzyme activity. The fluorogenic substrate Z-Gly-Pro-NH-Meq was also synthesized for comparison (Km = 10.35 10-6 mol l-1 at pH 7.0).

Synthesis and use of an aminoquinolinone derivative for the fluorometric determination of oxytocinase

1989 ◽  
Vol 54 (10) ◽  
pp. 2802-2808 ◽  
Author(s):  
Hana P. Mašková ◽  
George Kokotos ◽  
Chryssa Tzougraki ◽  
Tomislav Barth
Keyword(s):  
Organic Solvent ◽  
Human Serum ◽  
Enzyme Reaction ◽  
Natural Substrate ◽  
Fluorogenic Substrate ◽  
Chromogenic Substrates ◽  
Michaelis Constant ◽  
Fluorometric Determination ◽  
Optimum Ph

A new fluorogenic substrate for the determination of the activity of human serum oxytocinase-cystine aminopeptidase (EC 3.4.11.3), H-Cys(Bzl)-NH-Meq, has been synthesized. The affinity of H-Cys(Bzl)-NH-Meq to oxytocinase was by two orders higher than that of the usually employed chromogenic substrates. The Michaelis constant of oxytocinase for this substrate was within the range of the optimum pH (7.0–7.5) 2.3.10-6 mol l-1, i.e. in the region of the affinity of the natural substrate, oxytocin. The concentration of dimethylsulfoxide used for the solubilization of H-Cys(Byl)-NH-Meq (<0.4%) did not influence adversely the course of the enzyme reaction as in the case of chromogenic substrates, where the concentrations of the organic solvent exceeded 3%.

β-GLUCURONIDASE OF RABBIT POLYMORPHONUCLEAR LEUCOCYTES

1950 ◽  
Vol 28e (3) ◽  
pp. 69-79 ◽  
Author(s):  
R. J. Rossiter ◽  
Esther Wong
Keyword(s):  
Enzyme Activity ◽  
Blood Plasma ◽  
Substrate Concentration ◽  
Final Concentration ◽  
White Cell ◽  
Polymorphonuclear Leucocytes ◽  
Michaelis Constant ◽  
Glucuronidase Activity ◽  
Arsanilic Acid ◽  
Optimum Ph

Rabbit polymorphonuclear leucocytes contain an enzyme capable of hydrolyzing biosynthetic phenolphthalein mono-β-glucuronide. The concentration of the enzyme in the white cell is some 2000 times the concentration of the enzyme in the blood plasma. Under the conditions of study, the β-glucuronidase activity was proportional to the concentration of the enzyme. The effect of substrate concentration on the enzyme activity was studied and the Michaelis constant, Ks, determined. The course of the reaction was linear with time for the first 12 hr. and then fell off slightly during the next 12 hr. The optimum pH of the enzyme was 4.45 in either 0.2 M acetate or 0.2 M phthalate buffer. It was not inhibited by cyanide, azide, iodoacetate, fluoride, glycine, thiourea, urethane, arsanilic acid, acetophenone, o-cresol or m-cresol, in a final concentration of 0.01 M. The possible function of β-glucuronidase in rabbit polymorphonuclear leucocytes is discussed.

scholarly journals DETERMINATION OF HISTIDINE DECARBOXYLASE IN MICROGRAM SAMPLES OF TISSUE, PROPERTIES AND QUANTITATIVE HISTOCHEMISTRY OF THE ENZYME AND HISTAMINE IN THE RAT STOMACH

1967 ◽  
Vol 15 (6) ◽  
pp. 347-352 ◽  
Author(s):  
YOUNG S. KIM ◽  
DAVID GLICK
Keyword(s):  
Enzyme Activity ◽  
Histidine Decarboxylase ◽  
Preliminary Investigation ◽  
The Body ◽  
Rat Stomach ◽  
Michaelis Constant ◽  
Cell Region ◽  
Tissue Properties ◽  
Effects Of Ph

Properties of histidine decarboxylase of rat stomach were studied, including measurements of the Michaelis constant and effects of pH. Optimal conditions for determination of the enzyme were established. A method for this determination on microgram samples of tissue, e.g., microtome sections, that depends on fluorometric analysis of nanogram amounts of histamine, was elaborated and applied to a preliminary investigation of the quantitative histologic distribution of histidine decarboxylase in the body of the glandular stomach of the rat. A parallel study of the quantitative distribution of histamine was also made, and both it and the enzyme activity were predominantly concentrated in the chief cell region. From analysis of gross homogenates, the body of the stomach was found to have greater concentrations of histamine and enzyme activity than the antrum. The findings obtained were considered in relation to distribution of mast cells and acid secretion.

A Rapid Electrophoretic Method for the Determination of the Isoenzymes of Serum Lactate Dehydrogenase

1966 ◽  
Vol 12 (5) ◽  
pp. 308-313 ◽  
Author(s):  
Albert W Opher ◽  
Charles S Collier ◽  
Joseph M Miller
Keyword(s):  
Enzyme Activity ◽  
Lactate Dehydrogenase ◽  
Quantitative Determination ◽  
Serum Lactate ◽  
Serum Lactate Dehydrogenase ◽  
Electrophoretic Method ◽  
The Individual ◽  
Electrophoretic Procedure ◽  
Purple Formazan

Abstract A convenient electrophoretic procedure for the separation and quantitation of lactate dehydrogenase (LDH) isoenzymes is described. The system uses polyacetate Sepraphore III strips.* The areas of activity are shown by incubation with an LDH substrate combined with tetra-nitro-blue-tetrazolium. The reduction of the latter to the purple formazan is quantitatively related to the enzyme activity. Quantitative determination of the individual colored areas is performed by densitometry.

Sign in / Sign up

Export Citation Format

Share Document