INACTIVATION OF LUTEINIZING HORMONE RELEASING HORMONE BY RAT HYPOTHALAMIC L-CYSTINE ARYLAMIDASE

1975 ◽  
Vol 78 (4) ◽  
pp. 634-648 ◽  
Author(s):  
Herbert Kuhl ◽  
Hans-Dieter Taubert
Keyword(s):  
Inhibitory Effect ◽  
Supernatant Fraction ◽  
Luteinizing Hormone Releasing Hormone ◽  
Lysine Vasopressin ◽  
Hormone Effects ◽  
Rate Of Hydrolysis ◽  
Lh Rh ◽  
Hydrolysis Of ◽  
The Brain ◽  
Cystine Arylamidase

ABSTRACT The rate of hydrolysis of several aminoacyl-4-nitroanilides by rat hypothalamic arylamidases was investigated. The activity of these enzymes which were mainly found in the 105 000 × g supernatant fraction of homogenates of the hypothalamus and other parts of the brain was shown to depend upon the presence of metal ions and free thiol groups, and to be inhibited by puromycin. As previous investigations had shown that Cys-NA is an appropriate substrate for measuring hormone effects on hypothalamic arylamidases, L-cystine arylamidase and its interaction with various peptide hormones were examined in detail. It could conclusively be shown that this enzyme interacts particularly with oligopeptides. Its activity was competitively inhibited by TRF, oxytocin, lysine vasopressin, and LH-RH. It was also shown that the biological activity of LH-RH and its inhibitory effect on the hydrolysis of L-cystine-bis-p-nitroanilide decreased when it was incubated for various periods of time with the 105 000 × g supernatant of rat hypothalamus homogenate.

Polarographic study of hydrolysis of [8-lysine]vasopressin and its derivatives with blood serum of pregnant women

1980 ◽  
Vol 45 (4) ◽  
pp. 1099-1108 ◽  
Author(s):  
Mikuláš Chavko ◽  
Michal Bartík ◽  
Evžen Kasafírek
Keyword(s):  
Blood Serum ◽  
Pregnant Women ◽  
Degree Of Hydrolysis ◽  
Polarographic Study ◽  
Ph Optimum ◽  
Lysine Vasopressin ◽  
Rate Of Hydrolysis ◽  
Hydrolysis Of ◽  
Vasopressin Analogues

A polarographic study of the hydrolysis of [8-lysine]vasopressin and some hormonogens of the vasopressin series with the blood serum of women in the last week of pregnancy was studied. The dependence of hydrolysis on pH (pH optimum: 7.4-7.50, substrate concentration (Km 1.2 . 10-5M), pH stability and thermal stability were determined. The rate of hydrolysis of individual vasopressin analogues decreases in the order: [8-lysine]vasopressin > Nα-glycyl-prolyl[8-lysine]-vasopressin > Nα-leucyl-[8-lysine]vasopressin > Nα-alanyl-[8-lysine]vasopressin > Nα-phenyl alanyl-[8-lysine]vasopressin > Nα-diglycyl-[8-lysine]vasopressin > Nα-prolyl-[8-lysine]vasopressin > Nα-triglycyl-[8-lysine]vasopressin > Nα-sarcosyl-glycyl-[8-lysine]vasopressin. The degree of hydrolysis gradually increases to a multiple with the length of the pregnancy in consequence of the presence of oxytocine. However, vasopressin is also hydrolysed to a small extent with the enzymes from the blood sera of non-pregnant women. Under similar analytical conditions oxytocin was not hydrolysed with the sera of non-pregnant women and therefore oxytocin is a more suitable substrate than vasopressin for polarographic determination of serum oxytocinase.

scholarly journals Evidence for the regulation of guinea-pig heart microsomal phosphatidylcholine-hydrolysing phospholipase A1 by guanosine 5′-[γ-thio]triphosphate

1992 ◽  
Vol 288 (3) ◽  
pp. 965-968 ◽  
Author(s):  
K Badiani ◽  
X Lu ◽  
G Arthur
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Guinea Pig ◽  
Molecular Species ◽  
Inhibitory Effect ◽  
Phospholipase A1 ◽  
Guinea Pig Heart ◽  
Substrate Specificities ◽  
Rate Of Hydrolysis ◽  
Hydrolysis Of

We have recently characterized lysophospholipase A2 activities in guinea-pig heart microsomes and postulated that these enzymes act sequentially with phospholipases A1 to release fatty acids selectively from phosphatidylcholine (PC) and phosphatidylethanolamine, thus providing an alternative route to the phospholipase A2 mode of release. In a further investigation of the postulated pathway, we have characterized the PC-hydrolysing phospholipase A1 in guinea-pig heart microsomes. Our results show that the enzyme may have a preference for substrates with C16:0 over C18:0 at the sn-1 position. In addition, although the enzyme cleaves the sn-1 fatty acid, the rate of hydrolysis of PC substrates with C16:0 at the sn-1 position was influenced by the nature of the fatty acid at the sn-2 position. The order of decreasing preference was C18:2 > C20:4 = C18:1 > C16:0. The hydrolyses of the molecular species were differentially affected by heating at 60 degrees C. An investigation into the effect of nucleotides on the activity of the enzyme showed that guanosine 5′-[gamma-thio]triphosphate (GTP[S]) inhibited the hydrolysis of PC by phospholipase A1 activity, whereas GTP, guanosine 5′-[beta-thio]diphosphate (GDP[S]), GDP, ATP and adenosine 5′-[gamma-thio]triphosphate (ATP[S]) did not affect the activity. The inhibitory effect of GTP[S] on phospholipase A1 activity was blocked by preincubation with GDP[S]. A differential effect of GTP[S] on the hydrolysis of different molecular species was also observed. Taken together, the results of this study suggest the presence of more than one phospholipase A1 in the microsomes with different substrate specificities, which act sequentially with lysophospholipase A2 to release linoleic or arachidonic acid selectively from PC under resting conditions. Upon stimulation and activation of the G-protein, the release of fatty acids would be inhibited.

Differences between in vitro and in vivo degradation of LHRH by rat brain and other organs

1987 ◽  
Vol 253 (3) ◽  
pp. E317-E321 ◽  
Author(s):  
F. A. Carone ◽  
M. A. Stetler-Stevenson ◽  
V. May ◽  
A. LaBarbera ◽  
G. Flouret
Keyword(s):  
Degradation Products ◽  
Venous Blood ◽  
Simultaneous Action ◽  
Pituitary Cells ◽  
Time Intervals ◽  
Luteinizing Hormone Releasing Hormone ◽  
Hydrolysis Of ◽  
The Brain

Homogenates of brain, pituitary, liver, lung, ovary, and testes were incubated with [pyro Glu1-3,4-3H]luteinizing hormone-releasing hormone ([3H]LHRH), and the profiles of metabolites generated as a function of time were determined. After 5 min of incubation, 5 was the predominant metabolite in most homogenates. Although the profiles of metabolites varied at different time intervals, metabolites 2, 3, 4, and 5, and in some instances 7 and 9, appeared to form simultaneously and were detectable at 10 min. Neither metabolite 6 nor other larger metabolites formed initially as dominant degradation products. The findings suggest cleavage of LHRH by the simultaneous action of several endopeptidases. After a single vascular transit of [3H]LHRH, metabolites were determined in the venous blood of liver, lung, and brain of rats in vivo. There were no metabolites of [3H]LHRH in venous blood of liver and lung; however, metabolites 2-4 were present in venous blood of the brain. Incubation of rat anterior pituitary cells with [3H]LHRH yielded metabolites 1-4 but not metabolites 5 or 9 as in homogenates. Incubation of [3H]LHRH with porcine follicular granulosa cells resulted in the generation of metabolites 2-7 and 9, similar to the profile in homogenates. Thus, since homogenates contain enzymes of disrupted cells, they do not always reflect mechanisms for in vivo hydrolysis of circulating LHRH. Brain degraded 12.1% of LHRH during a single vascular transit and may account for substantial degradation of the circulating hormone.

THE PRESENCE OF PEPTIDASES IN THE RAT HYPOTHALAMUS INACTIVATING LUTEINIZING HORMONE RELEASING HORMONE (LH-RH)

1974 ◽  
Vol 77 (3) ◽  
pp. 435-442 ◽  
Author(s):  
E. C. Griffiths ◽  
K. C. Hooper ◽  
S. L. Jeffcoate ◽  
D. T. Holland
Keyword(s):  
Luteinizing Hormone ◽  
Reproductive Function ◽  
Assay Method ◽  
Supernatant Fraction ◽  
Peptidase Activity ◽  
Female Rat ◽  
Luteinizing Hormone Releasing Hormone ◽  
Releasing Hormone ◽  
Rat Hypothalamus ◽  
Lh Rh

ABSTRACT The presence of peptidases in the rat hypothalamus inactivating luteinizing hormone-releasing hormone (LH-RH) has previously been demonstrated using an indirect assay method. With the development of a sensitive and specific radioimmunoassay for the releasing hormone, this technique was employed in the study of the peptidases inactivating LH-RH. It was found that supernatant fractions from both male and female rat hypothalami rapidly inactivated LH-RH, and that the peptidase activity of the supernatant fraction was higher in male than in female animals though the particulate fraction's activity was about the same in both sexes. Peptidase activity was also considerably greater in the supernatant than in the particulate fraction. These results confirm that the hypothalamus contains peptidases capable of inactivating LH-RH and give a direct indication that the enzymes may be involved in the central nervous system's control of reproductive function.

USE OF YEAST BETA-GALACTOSIDASE IN MILK AND MILK PRODUCTS1

1971 ◽  
Vol 34 (6) ◽  
pp. 294-299 ◽  
Author(s):  
W. L. Wendorff ◽  
C. H. Amundson ◽  
N. F. Olson ◽  
J. C. Garver
Keyword(s):  
Maximum Rate ◽  
Inhibitory Effect ◽  
Potassium Sulfate ◽  
Factors Affecting ◽  
Milk Products ◽  
Dry Milk ◽  
Rate Of Hydrolysis ◽  
Milk Solids ◽  
Hydrolysis Of ◽  
Beta Galactosidase

Experiments were carried out to study factors affecting the enzyme activity of β-galactosidase of Saccharomyces fragilis NRRL Y-1109 in milk products. Both the type of lactose-containing substrates and their method of preparation greatly affected β-galactosidase activity. Lactose in an aqueous solution was hydrolyzed more easily than it was in milk products. Of milk products tested, whey was the best substrate for the enzyme. Milk solids, other than lactose, exhibited some inhibitory effect on hydrolysis of lactose by the S. fragilis β-galactosidase. The maximum rate of hydrolysis in milk products was obtained when milk or whey was fortified with 0.1 M potassium sulfate and 10−4 M manganese chloride. Nonfat dry milk and whey powders, in which portions of the lactose were hydrolyzed to simple sugars, were prepared. These products were of good flavor, appearance, and stability.

scholarly journals Immunohistochemical localization of hypothalamic hormones.

1976 ◽  
Vol 24 (7) ◽  
pp. 864-871 ◽  
Author(s):  
G Pelletier ◽  
R Leclerc ◽  
D Dubé
Keyword(s):  
Endocrine Cells ◽  
Area Postrema ◽  
Secretory Granules ◽  
Subcommissural Organ ◽  
Immunohistochemical Localization ◽  
Nerve Endings ◽  
Immunoperoxidase Technique ◽  
Luteinizing Hormone Releasing Hormone ◽  
Lh Rh ◽  
The Brain

Using an immunoperoxidase technique at the light and electron microscope levels, the localization of somatostatin and luteinizing hormone-releasing hormone (LH-RH) has been performed in the brain, as well as the pancreas and stomach. The following generalizations may be drawn on the localization of LH-RH and somatostatin. (a) LH-RH and somatostatin are contained in axons and nerve endings of the median eminence and organum vasculosum of the lamina terminalis. (b) LH-RH and somatostatin are present in different nerve endings. (c) In the subcommissural organ, subfornical organ and area postrema, the two neurohormones are present in the cytoplasm of ependyman and subependymal cells. (d) In the endocrine pancreas and gastric mucosa, somatostatin is localized within the secretory granules of specific endocrine cells.

The Effects of Amines on the Esterase Activities of Thrombin and Thrombokinase and on Several Clotting Tests

1974 ◽  
Vol 31 (02) ◽  
pp. 309-318
Author(s):  
Phyllis S Roberts ◽  
Raphael M Ottenbrite ◽  
Patricia B Fleming ◽  
James Wigand
Keyword(s):  
Choline Chloride ◽  
Polymerization Process ◽  
Ammonium Bromide ◽  
Bovine Thrombin ◽  
One Stage ◽  
Human Proteins ◽  
Rate Of Hydrolysis ◽  
The One ◽  
Hydrolysis Of Esters ◽  
Hydrolysis Of

Summary1. Choline chloride, 0.1 M (in 0.25 M Tris. HCl buffer, pH 7.4 or 8.0, 37°), doubles the rate of hydrolysis of TAME by bovine thrombokinase but has no effect on the hydrolysis of this ester by either human or bovine thrombin. Only when 1.0 M or more choline chloride is present is the hydrolysis of BAME by thrombokinase or thrombin weakly inhibited. Evidence is presented that shows that these effects are due to the quaternary amine group.2. Tetramethyl ammonium bromide or chloride has about the same effects on the hydrolysis of esters by these enzymes as does choline chloride but tetra-ethyl, -n.propyl and -n.butyl ammonium bromides (0.1 M) are stronger accelerators of the thrombokinase-TAME reaction and they also accelerate, but to a lesser degree, the thrombin-TAME reaction. In addition, they inhibit the hydrolysis of BAME by both enzymes. Their effects on these reactions, however, do not follow any regular order. The tetraethyl compound is the strongest accelerator of the thrombokinase-TAME reaction but the tetra-ethyl and -butyl compounds are the strongest accelerators of the thrombin-TAME reaction. The ethyl and propyl compounds are the best (although weak) inhibitors of the thrombokinase-BAME and the propyl compound of the thrombin-BAME reactions.3. Tetra-methyl, -ethyl, -n.propyl and -n.butyl ammonium bromides (0.01 M) inhibit the clotting of fibrinogen by thrombin (bovine and human proteins) at pH 7.4, imidazole or pH 6.1, phosphate buffers and they also inhibit, but to a lesser degree, a modified one-stage prothrombin test. In all cases the inhibition increases regularly as the size of the alkyl group increases from methyl to butyl. Only the ethyl com pound (0.025 M but not 0.01 M), however, significantly inhibits the polymerization of bovine fibrin monomers. It was concluded that inhibition of the fibrinogen-thrombin and the one-stage tests by the quaternary amines is not due to any effect of the com pounds on the polymerization process but probably due to inhibition of thrombin’s action on fibrinogen by the quaternary amines.

INCORPORATION OF IODIDE INTO ORGANIC COMPOUNDS IN THE GUINEA PIG THYROID

1963 ◽  
Vol 43 (1) ◽  
pp. 110-118 ◽  
Author(s):  
R. Ekholm ◽  
T. Zelander ◽  
P.-S. Agrell
Keyword(s):  
Guinea Pig ◽  
Protein Binding ◽  
Organic Compounds ◽  
Guinea Pigs ◽  
Microsomal Fraction ◽  
Thyroid Tissue ◽  
Particulate Fraction ◽  
Supernatant Fraction ◽  
Hydrolysis Of

ABSTRACT Guinea pigs, kept on a iodine-sufficient diet, were injected with Na131I and the thyroids excised from 45 seconds to 5 days later. The thyroid tissue was homogenized and separated into a combined nuclear-mitochondrial-microsomal fraction and a supernatant fraction by centrifugation at 140 000 g for one hour. Protein bound 131iodine (PB131I) and free 131iodide were determined in the fractions and the PB131I was analysed for monoiodotyrosine (MIT), diiodotyrosine (DIT) and thyroxine after hydrolysis of PB131I. As early as only 20 minutes after the Na131I-injection almost 100% of the particulate fraction 131I was protein bound. In the supernatant fraction the protein binding was somewhat less rapid and PB131I values above 90% of total supernatant 131I were not found until 3 hours after the injection. In all experiments the total amount of PB131I was higher in the supernatant than in the corresponding particulate fraction. The ratio between supernatant PB131I and pellet PB131I was lower in experiments up to 3 minutes and from 2 to 5 days than in experiments of 6 minutes to 20 hours. Hydrolysis of PB131I yielded, even in the shortest experiments, both MIT and DIT. The DIT/MIT ratio was lower in the experiments up to 2 hours than in those of 3 hours and over.

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