scholarly journals The hydrolysis of phosphatidylinositol monolayers at an air/water interface by the calcium-ion-dependent phosphatidylinositol phosphodiesterase of pig brain

1981 ◽  
Vol 193 (2) ◽  
pp. 607-614 ◽  
Author(s):  
K Hirasawa ◽  
R F Irvine ◽  
R M C Dawson
Keyword(s):  
Oleyl Alcohol ◽  
Enzymic Activity ◽  
Lag Phase ◽  
Water Interface ◽  
Inhibitory Effects ◽  
Pig Brain ◽  
Air Water Interface ◽  
Rate Of Hydrolysis ◽  
Hydrolysis Of

1. The activity of Ca2+-dependent phosphatidylinositol phosphodiesterase (EC 3.1.4.10) of pig brain against [32P]phosphatidylinositol monolayers at an air/water interface has been measured. As the monolayer pressure was increased a sharp cut-off of enzymic hydrolysis occurred at 33 × 10(-3) N/m. 2. The addition of either phosphatidic acid, phosphatidylglycerol or oleyl alcohol increased the film pressure at which cut off occurred, as well as increasing the rate of hydrolysis at lower pressures. 3. The rate of hydrolysis, but not the cut-off pressure, was markedly increased by oleic acid and slightly increased by phosphatidylethanolamine. 4. Phosphatidylcholine, palmitoylcholine and octadecylamine decreased the cut-off pressure, as well as the enzymic activity below this pressure. 5. Stearic acid and stearyl alcohol had no effect on either the cut-off pressure or the activity. 6. All activators decreased the length of the lag phase before enzyme activity began, and phosphatidylcholine increased it. 7. These results are compared with the stimulatory and inhibitory effects of various amphiphiles observed previously with phosphatidylinositol dispersions [Irvine, Hemington & Dawson (1979) Eur. J. Biochem. 99, 525-530], and their possible relevance to the control of the phosphatidylinositol phosphodiesterase in vivo are discussed.

Studies on the mechanism of acute inhibition of thyroglobulin hydrolysis by iodine

1985 ◽  
Vol 108 (4) ◽  
pp. 511-517 ◽  
Author(s):  
Nandalal Bagchi ◽  
Birdie Shivers ◽  
Thomas R. Brown
Keyword(s):  
Time Course ◽  
Period 2 ◽  
Iodotyrosine Deiodinase ◽  
Rate Of Hydrolysis ◽  
Underlying Mechanisms ◽  
Excess Iodide ◽  
Sites Of Action ◽  
Hydrolysis Of

Abstract. Iodine in excess is known to acutely inhibit thyroidal secretion. In the present study we have characterized the time course of the iodine effect in vitro and investigated the underlying mechanisms. Labelled thyroid glands were cultured in vitro in medium containing mononitrotyrosine, an inhibitor of iodotyrosine deiodinase. The rate of hydrolysis of labelled thyroglobulin was measured as the proportion of labelled iodotyrosines and iodothyronines recovered at the end of culture and was used as an index of thyroidal secretion. Thyrotrophin (TSH) administered in vivo acutely stimulated the rate of thyroglobulin hydrolysis. Addition of Nal to the culture medium acutely inhibited both basal and TSH-stimulated thyroglobulin hydrolysis. The effect of iodide was demonstrable after 2 h, maximal after 6 h and was not reversible upon removal of iodide. Iodide abolished the dibutyryl cAMP induced stimulation of thyroglobulin hydrolysis. Iodide required organic binding of iodine for its effect but new protein or RNA synthesis was not necessary. The inhibitory effects of iodide and lysosomotrophic agents such as NH4C1 and chloroquin on thyroglobulin hydrolysis were additive suggesting different sites of action. Iodide added in vitro altered the distribution of label in prelabelled thyroglobulin in a way that suggested increased coupling in the thyroglobulin molecule. These data indicate that 1) the iodide effect occurs progressively over a 6 h period, 2) continued presence of iodide is not necessary once the inhibition is established, 3) iodide exerts its action primarily at a post cAMP, prelysosomal site and 4) the effect requires organic binding of iodine, but not new RNA or protein synthesis. Our data are consistent with the hypothesis that excess iodide acutely inhibits thyroglobulin hydrolysis by increasing the resistance of thyroglobulin to proteolytic degradation through increased iodination and coupling.

Repeated thyrotrophin stimulation of thyroid secretion: lack of refractoriness in vivo

1985 ◽  
Vol 106 (2) ◽  
pp. 153-157
Author(s):  
N. Bagchi ◽  
T. R. Brown
Keyword(s):  
Adenylate Cyclase ◽  
Thyroid Tissue ◽  
Control Group ◽  
Cyclic Amp Accumulation ◽  
Thyroid Glands ◽  
Rate Of Hydrolysis ◽  
Thyroid Secretion ◽  
Hydrolysis Of

ABSTRACT It has been reported that prior exposure of thyroid tissue to TSH in vitro induces a state of refractoriness to new challenges of the hormone. We have investigated the effect of repeated TSH treatment on thyroid secretion to determine whether such refractoriness exists in vivo. The rate of thyroid secretion was estimated by measuring the rate of hydrolysis of labelled thyroglobulin from mouse thyroid glands in vitro. The thyroid glands were labelled in vivo with 131I and then cultured for 20 h in the presence of mononitrotyrosine, an inhibitor of iodotyrosine deiodinase. The rate of hydrolysis of labelled thyroglobulin was measured as the percentage of radioactivity released as free iodotyrosines and iodothyronines into the gland and the medium at the end of incubation. Thyrotrophin was administered in vivo at hourly intervals for 2–4 injections. The corresponding control group received saline injections every hour except for the last injection when they received TSH. The peak rates of thyroglobulin hydrolysis, measured 2 h following the last injection, were similar in animals receiving two, three or four TSH injections and were not different from those in the control groups. Serum tri-iodothyronine and thyroxine concentrations 2 h after the last injection were higher in the groups receiving multiple TSH injections. Thyroidal cyclic AMP accumulation in response to TSH was markedly depressed in the group receiving multiple injections compared with the group receiving a single injection of TSH in vivo. These data indicate that (1) the stimulatory effect of TSH on thyroidal secretion is not diminished by prior administration of the hormone in vivo, (2) repeated TSH administrations in vivo cause refractoriness of the adenylate cyclase response to TSH and (3) a dichotomy exists between the secretory response and the adenylate cyclase response to repeated administrations of TSH. J. Endocr. (1985) 106, 153–157

scholarly journals Observations on the action of limit dextrinases on amylopectin-like polysaccharides

1973 ◽  
Vol 133 (2) ◽  
pp. 413-416 ◽  
Author(s):  
G. Dunn ◽  
D. G. Hardie ◽  
D. J. Manners
Keyword(s):  
Rate Of Hydrolysis ◽  
Limit Dextrinase ◽  
Hydrolysis Of

The rate of hydrolysis of amylopectin by three different limit dextrinase preparations is only about 15–23% of that of amylopectin β-limit dextrin under similar conditions. On dilution of the enzymes there was no change in specificity. The factors controlling the specificity of the enzyme and the possible significance in vivo of the results are discussed.

A new succinylcholine-based assay of plasma cholinesterase.

1984 ◽  
Vol 30 (2) ◽  
pp. 192-195 ◽  
Author(s):  
M H Abernethy ◽  
P M George ◽  
V E Melton
Keyword(s):  
Hydrogen Peroxide ◽  
Enzyme Activity ◽  
Plasma Cholinesterase ◽  
Choline Oxidase ◽  
Rate Of Hydrolysis ◽  
The Mean ◽  
Variant Enzyme ◽  
Hydrolysis Of

Abstract We describe a new method for measuring the in vitro rate of hydrolysis of the muscle relaxant succinylcholine. This substrate is hydrolyzed by plasma cholinesterase (EC 3.1.1.8). The resulting choline is determined by measuring the hydrogen peroxide formed on its oxidation by choline oxidase (EC 1.1.3.17). This is done by use of phenol and aminoantipyrine coupled to peroxidase, and yields an intense chromophore, Amax 500 nm. The assay requires 0.1 mL of plasma, and is precise and specific. The CV was 2.7% within run, 7.3% between run. For the usual (U variant) enzyme the Km is 53 mumol/L. Enzyme activity is removed by anticholinesterase antiserum, and is inhibited by dibucaine with a Ki of 2 mumol/L. Ten samples can be assayed in duplicate in an hour. This method is suited to routine use in any laboratory that has a simple spectrophotometer. The mean activity in 11 individuals with the cholinesterase phenotype UU was 105 U/L, for seven UA heterozygotes 61 U/L, and for three AA homozygotes 4 U/L. To the extent allowed by extrapolation from in vitro to in vivo results, this method should increase diagnostic accuracy and may directly predict duration of succinylcholine-induced apnea.

Preferential in vivo accumulation of sn-2,3-diacylglycerols in postheparin plasma of rats

1977 ◽  
Vol 55 (10) ◽  
pp. 1075-1081 ◽  
Author(s):  
N. Morley ◽  
A. Kuksis ◽  
A. G. D. Hoffman ◽  
G. Kakis
Keyword(s):  
Portal Vein ◽  
Lipoprotein Lipase ◽  
Hepatic Lipase ◽  
Water Interface ◽  
Substrate Complex ◽  
Enzyme Substrate ◽  
Oil Water ◽  
Hydrolysis Of

The stereochemical course of in vivo hydrolysis of triacylglycerols by lipoprotein lipase was investigated by determining the structure of diacylglycerol intermediates in postheparin plasma of rats which had been fed [3H]glycerol-labeled Intralipid 2 h before an injection of heparin or had been given an injection of a mixture of [3H]glycerol-Intralipid and heparin. During the clearance of both the natural chylomicrons and the artificial emulsion, sn-2,3-diacylglycerols (60–80%) were found to be the dominant enantiomers. Similar results were obtained when the contribution of the hepatic lipase was altered, either by tying off the mesentery artery and portal vein before injection of heparin, or by injection of heparin directly into the portal vein. These findings are consistent with a preferential release of the acyl group from the sn-1 position of the triacylglycerol molecule as demonstrated previously in vitro. A preferential orientation of the substrate in the enzyme–substrate complex or at the oil–water interface is discussed as a possible basis for these effects.

scholarly journals Degradation of pyrene-labelled phospholipids by lysosomal phospholipases in vitro. Dependence of degradation on the length and position of the labelled and unlabelled acyl chains

1996 ◽  
Vol 315 (3) ◽  
pp. 947-952 ◽  
Author(s):  
S Lusa ◽  
M Myllarniemi ◽  
K Volmonen ◽  
M Vauhkonen ◽  
P Somerharju
Keyword(s):  
Fatty Acid ◽  
Acyl Chain ◽  
Upward Movement ◽  
Lysosomal Degradation ◽  
Acyl Chains ◽  
Rate Of Hydrolysis ◽  
Carboxy Terminal ◽  
Hydrolysis Of

The hydrolysis of pyrenylacyl phosphatidylcholines (PyrnPCs) (n indicates the number of aliphatic carbons in the pyrene-chain) by crude lysosomal phospholipases in vitro was investigated. PyrnPCs consist of several sets in which the length of the pyrene-labelled or the unlabelled acyl chain, linked to the sn-1 or sn-2 position, was systematically varied. Lysophosphatidylcholine and fatty acid were the only fluorescent breakdown products detected, thus indicating that PyrnPCs were degraded by A-type phospholipases and lysophospholipases. Of these, mainly A1-type phospholipases appear to be involved, as determined from the relative amounts of labelled fatty acid and lysolipid released from the positional isomers. Based on the effects of the length and position of the pyrene-labelled and unlabelled chains it is suggested that (1) the lysosomal A-type phospholipases acting on PyrnPCs recognize the carboxy-terminal part of the lipid acyl chains and (2) the relevant part of the binding site is relatively narrow. Thus phospholipids with added bulk in the corresponding region, such as those that are peroxidized and polymerized, may not be good substrates for the lysosomal phospholipases mentioned. The impaired hydrolysis of the most hydrophobic PyrnPCs indicates that lysosomal phospholipases may not be able to penetrate significantly into the substrate interphase, but upward movement of the lipid may be required for efficient hydrolysis. Finally, the rate of hydrolysis of many pyrenyl derivatives was found to be comparable to that of a natural phosphatidylcholine species, both in micelles and in lipoprotein particles, indicating that these derivatives can be used as faithful reporters of lysosomal degradation of natural lipids in vivo and in vitro.

The Surface Chemistry of Proteins. III. The Rigidity of Adsorbed Films at an Air-Water Interface

1952 ◽  
Vol 5 (1) ◽  
pp. 189
Author(s):  
CWN Cumper ◽  
AE Alexander
Keyword(s):  
Surface Chemistry ◽  
Oleyl Alcohol ◽  
Water Interface ◽  
Adsorbed Films ◽  
Alcohol Water ◽  
Air Water Interface ◽  
Oil Water ◽  
Petrol Ether ◽  
Surface Rigidity

The results presented in Part II of this series on the adsorption of the proteins, β-globulin, pepsin, and insulin at a white oil, 5 per cent, oleyl alcohol-water interface have been extended to the petrol ether-water and air-water interfaces by studying the surface rigidity of the films formed. The previous picture of the process of adsorption has been generally confirmed but there are certain differences between the oil-water and air-water interfaces which are discussed.

scholarly journals Regulation of NAD+ glycohydrolase activity by NAD+-dependent auto-ADP-ribosylation

1996 ◽  
Vol 318 (3) ◽  
pp. 903-908 ◽  
Author(s):  
Myung-Kwan HAN ◽  
Ji-Young LEE ◽  
Yee-Sook CHO ◽  
Young M SONG ◽  
Nyeon-Hyoung AN ◽  
...  
Keyword(s):  
Enzyme Activity ◽  
Physiological Role ◽  
Enzymic Activity ◽  
Adp Ribosylation ◽  
Nad Glycohydrolase ◽  
Intact Rabbit ◽  
Hydrolysis Of

NAD+ glycohydrolase (NADase; EC 3.2.2.5) is an enzyme that catalyses hydrolysis of NAD+ to produce ADP-ribose and nicotinamide. Its physiological role and the regulation of its enzymic activity have not been fully elucidated. In the present study, the mechanism of self-inactivation of NADase by its substrate, NAD+, was investigated by using intact rabbit erythrocytes and purified NADase. Our results suggest that inactivation of NADase was due an auto-ADP-ribosylation reaction. ADP-ribosylated NADase of rabbit erythrocytes was deADP-ribosylated when incubated without NAD+, and thus enzyme activity was simultaneously restored. These findings suggest that reversible auto-ADP-ribosylation of NADase might regulate the enzyme's activity in vivo.

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