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Enzyme
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Query: EC:3.4.21.4 (
trypsin
)
42,187
document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)
Two highly purified sarcoplasmic reticulum membrane fractiones differing in their sensitivities to the uncoupling action of
caffeine
were isolated from white skeletal muscles of the rabbit. The main protein component of both fractions is a catalytical polypeptide of Ca2+-dependent ATPase. Treatment of the
caffeine
-sensitive reticular fraction by
trypsin
or DTNB completely removes the effect of
caffeine
. It was found that similar effects on the
caffeine
-sensitive reticular fraction are exerted by bemegride, camphor, ethymizole and cordiamine. Isolation of Ca2+-dependent ATPase from both reticular fractions and reconstruction of Ca2+-transporting vesicles were carried out. Ca2+ transport by the vesicles enriched by ATPase from the
caffeine
-sensitive reticular fraction is uncoupled under the effect of
caffeine
; however,
caffeine
has no effect on the vesicles enriched by
caffeine
-insensitive reticular ATPase. The molecular weight of
caffeine
-sensitive and
caffeine
-insensitive ATPases determined in the presence of sedium dodecyl sulfate are found to be identical. Electrophoresis in the presence of digitonin revealed different electrophoretic behaviour of the two forms of ATPase.
...
PMID:[Two forms of Ca2+-dependent ATPase from sarcoplasmic reticulum]. 15 30
The effects of local anesthetic agents (lidocaine, procaine, cocaine) and diphenylhydantoin (DPH) were studied on the slow electrical responses induced by isoproterenol or
caffeine
in cardiac muscle preparations rendered inexcitable by tetrodotoxin (TTX) or by partial depolarization with elevated K+ (26 mM). In such inexcitable cells, we previously demonstrated that addition of some positive inotropic agents, such as catecholamines, histamine, and methylxanthines, rapidly increase the number of available slow Ca2+--Na+ channels, thus allowing slowly rising electrical responses resembling the plateau component of the cardiac action potential. In embryonic chick (16-20-day-old) myocardial cells (ventricular) studied as intact perfused hearts or as reaggregated cell cultures of
trypsin
-dispersed cells, high concentrations (10-(3) M) of all of the above drugs blocked the induced slow responses with their associated contractions; low concentrations (10-(5) M) of these agents reduced the maximal rate of rise (+Vmax) of the slow responses and depressed the contractions. For comparison with their effects on the slow response, the actions of these drugs on the normal action potential were also studied. As with the slow response, all of these drugs depressed the rate of rise of the action potential (10-(4) M) or blocked it at higher concentrations (10-(3) M); in contrast, low concentrations (10-(5) M) of lidocaine and DPH increased +V max. These findings suggest that local anesthetics, which interact with the lipid phase of the cell membrane, lead to blockade of the slow Ca2+--Na+ channels as well as of the fast Na+ channels in the myocardial sarcolemma.
...
PMID:Local anesthetic blockade of Ca2+ -mediated action potentials in cardiac muscle. 99 32
Fibrinogen degradation products (FDP) from bovine fibrinogen were obtained in vitro by
trypsin
digestion for 3 and 120 min, respectively. FDP-120 min applied i.p. and i.v.c. stimulated the CNS, while FDP-3 min did not. The FDP increased the potency of amphetamine,
caffeine
, thiopental, and chlorpromazine by enhancing their level in the brain. Peptides from 120 min digestion were more effective. A possible mechanism of observed changes has been discussed.
...
PMID:Influence of fibrinogen degradation products (FDP) on the central nervous system and on the effects of centrally acting drugs. 115 9
We studied the effect of two peptides, the intracellular Ca2(+)-mobilizing caerulein and the cAMP-mediated secretin, and also the phosphodiesterase inhibitor
caffeine
on pancreatic secretion and growth in newborn rats. To investigate the secretory response of these substances, and to construct dose-response curves 10-day-old conscious rats were given subcutaneously a single injection of caerulein, secretin,
caffeine
or the combination of these compounds (caerulein + secretin, caerulein +
caffeine
). Fifteen min after the injection pancreatic specific
trypsin
activities were measured in order to estimate depletion of enzymes from the pancreas. To study the pancreatic growth--promoting effect of these substances, newborn rats were treated three times daily for 10 days from the day of birth, using the same experimental groups as described above. Caerulein stimulated both enzyme depletion and pancreatic growth. Secretin stimulated enzyme depletion and increased the trophic effects of caerulein on the pancreas.
Caffeine
alone or in combination with caerulein did not affect pancreatic enzyme depletion and growth.
...
PMID:Secretin potentiates, caffeine does not affect caerulein--stimulated pancreatic enzyme depletion and growth in newborn rats. 169
An in vitro endosome fusion assay using Dictyostelium discoideum is described. The method requires endocytosis of anti-dinitrophenol (DNP) IgG or DNP-derivitized beta-glucuronidase into two sets of cells. After homogenizing the cells, the vesicles were mixed, and fusion was measured by quantitating immune complex formation between DNP-beta-glucuronidase and anti-DNP IgG. Fusion was dependent upon ATP, temperature, pH, ionic strength, and cytosol and sensitive to detergent, dilution,
trypsin
, N-ethylmaleimide, and guanosine 5'-3-O-(thio)triphosphate. Although weak bases, ionophores, hadacidin, [ethylenebis(oxyethylenenitrilo)]tetraacetic acid, and
caffeine
inhibit endocytosis in vivo, these reagents had no affect on in vitro endosome fusion. Comparison of Dictyostelium with mammalian cells showed differences in the temperature, pH, and salt requirements for fusion, possibly reflecting differences in the life-styles of various cell types. Like mammalian cells, Dictyostelium required GTP-binding protein(s) and an N-ethylmaleimide-sensitive factor for endosome fusion. Thus, the mechanism driving endosome fusion may have been conserved throughout evolution. Electron microscopic studies confirmed in vitro endosome fusion and revealed endosomes were being engulfed by other endosomes, resulting in formation of multivesicular elements (i.e. autophagic vesicles). This system may be useful for characterizing mutations, evolution, and developmental regulation along the endocytic pathway.
...
PMID:Characterization of endosome-endosome fusion in a cell-free system using Dictyostelium discoideum. 173 Jul 25
In this study, we found that sodium selenite was potent in inducing contracture of the mouse diaphragm. The possible mechanism of action of selenite was investigated. Contracture was induced by a direct action of selenite on the muscle membrane rather than that selenite enhanced transmitter release from the motor nerve terminals, since denervation, d-tubocurarine and tetrodotoxin did not inhibit the selenite-induced contracture. Although selenite decreased both the membrane potential and the amplitude of the muscle action potential, neither high K+ nor glycerol treatment, which closed the transverse tubule, reduced the selenite-induced contracture, suggesting that depolarization of the muscle membrane was not essential for the induction of the contracture. EGTA (1-50 mM) inhibited the selenite-induced contracture in a concentration-dependent manner. In contrast, varying the external Ca2+ concentrations from 10(-3) to 10 mM or raising Mg2+ concentration to 10 mM did not affect the contracture. Similarly, the contracture induced by
caffeine
was not affected by lowering the external Ca2+ concentration to 10(-3) mM but was completely inhibited by 30 mM EGTA. Selenite pretreatment markedly potentiated the
caffeine
contracture and prolonged treatment with
caffeine
inhibited the selenite contracture. All of these findings suggest that the selenite contracture was not dependent on external Ca2+ but was induced by the release of Ca2+ from internal membranes such as the sarcoplasmic reticulum. Pretreatment with
trypsin
, glutathione or cyanide blocked the selenite-evoked contracture. Therefore, we postulate that the selenite-induced contracture was induced by the initial binding of selenite to the sulfhydryl groups of the muscle membrane, which then triggered the release of Ca2+ from internal membranes such as the sarcoplasmic reticulum.
...
PMID:Studies on the contracture of the mouse diaphragm induced by sodium selenite. 250 65
Mouse diaphragm contractures induced by Cu2+,
caffeine
and selenite were studied comparatively. Both Cu2+- and
caffeine
-contractures were produced rapidly and relaxed spontaneously; the selenite-contracture occurred after a latent period of about 45 min and lasted for more than 3 hr. All contractures were myogenic, since neither d-tubocurarine nor tetrodotoxin prevented them. The susceptibility of these contractures to the depletion and replenishment of Ca2+ differed: the Cu2+-contracture increased proportionally with rising extracellular Ca2+ concentrations ranging from 2.5 to 12.5 mM and were abolished by 5 mM EGTA.
Caffeine
- and selenite-contractures were not affected by changes in extracellular Ca2+ concentration. The
caffeine
-contracture was abolished by EGTA in high concentration (30 mM) and the selenite-contracture was inhibited by 50 mM EGTA. After removal of Ca2+ with 5 mM EGTA, followed by replacement with 2.5 mM Ca2+ for 1 min, the Cu2(+)-contracture was fully restored.
Caffeine
- and selenite-contractures were restored only after a longer period (10-20 min) of re-exposure to Ca2+. These findings suggest that the Cu2(+)-contracture is dependent on external Ca2+ and probably caused by an increasing Ca2+ entry through sarcolemma.
Caffeine
- and selenite-contractures apparently result from internal Ca2+ release by sarcoplasmic reticulum. Substitution of either Sr2+ or Co2+ for Ca2+ fully supports the Cu2(+)-contracture. 45Ca2+ uptake and calcium content of the diaphragm were markedly increased by Cu2+ but not by selenite. Furthermore, the Cu2(+)-contracture was inhibited by exposing the outer membrane to
trypsin
, phospholipase C or saponin. The selenite-contracture was inhibited only by
trypsin
. The
caffeine
-contracture was unaffected by these treatments. These results support the notion that the Cu2(+)-contracture is induced by an increased entry of Ca2+ through the outer membrane. Cu2(+)-,
caffeine
- and selenite-contractures were respectively abolished, potentiated and unaffected by chronic denervation of the diaphragm. This and the other findings provide evidence that Cu2(+)-,
caffeine
- and selenite-contractures are induced in mouse diaphragm muscle via different sites of action.
...
PMID:Studies on contractures induced in mouse diaphragm by caffeine and cupric and selenite ions. 251 19
The subunit structure of the rabbit skeletal muscle ryanodine receptor-Ca2+ release channel complex was examined following solubilization of heavy sarcoplasmic reticulum membranes in two zwitterionic detergents, 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acid (Chaps) and Zwittergent 3-14. High and low affinity [3H]ryanodine binding was retained upon solubilization of the complex in Chaps but was lost in Zwittergent 3-14. The purified complex migrated as a single peak with an apparent sedimentation coefficient of approximately 30 and approximately 9 S upon density gradient centrifugation and with isoelectric points of 3.7 and 3.9 upon two-dimensional gel electrophoresis in Chaps and Zwittergent 3-14, respectively. Electron microscopy of negatively stained samples indicated that the distinct four-leaf clover structure of the ryanodine receptor observed in Chaps disappeared following Zwittergent treatment of the 30 S complex and instead showed smaller, round particles. Ferguson plot analysis following sodium dodecyl sulfate-polyacrylamide gel electrophoresis of partial and fully cross-linked and incompletely denatured complexes suggested a stoichiometry of four Mr approximately 400,000 peptides/30 S ryanodine receptor oligomer. [3H]Ryanodine binding to the membrane-bound receptor in 50 microM--1 mM free Ca2+ revealed the presence of both high affinity (KD = 8 nM, Hill coefficient (nH) = 0.9) and low affinity (nH approximately 0.45) sites with a ratio of 1:3. Reduction in free Ca2+ to less than or equal to 0.1 microM or
trypsin
digestion of the membranes resulted in loss of high affinity but not low affinity ryanodine binding (Hill KD = 5,000 nM, nH = 0.9). Addition of 20 mM
caffeine
to the nanomolar Ca2+ medium decreased the Hill KD to 1,000 nM without changing the Hill coefficient. Occupation of the low affinity sites altered the rate of [3H]ryanodine dissociation from the high affinity sites. Single channel recordings of the purified ryanodine receptor channel incorporated into planar lipid bilayers also indicated the existence of high and low affinity sites for ryanodine, occupation of which resulted in formation of a subconducting and completely closed state of the channel, respectively. These results are compatible with a subunit structural model of the 30 S ryanodine receptor-Ca2+ release channel complex which comprises a homotetramer of negatively charged and allosterically coupled polypeptides of Mr approximately 400,000.
...
PMID:The ryanodine receptor-Ca2+ release channel complex of skeletal muscle sarcoplasmic reticulum. Evidence for a cooperatively coupled, negatively charged homotetramer. 255 Apr 60
Coffee consumption has been associated with pancreatic disorders, but the mechanisms involved remain to be elucidated. This investigation examines the effects of
caffeine
consumption on the structure and function of the exocrine pancreas. Groups of rats, fed ad libitum commercial laboratory diet, were given drinking water which contained either
caffeine
(0.09 mg/ml) or nothing at all. The rats were allowed drink ad libitum and were killed 6 weeks later. Final body and pancreatic weights were not significantly different between the groups at the end of the experimental period. Although no ultrastructural effects of
caffeine
on the pancreas were observed, amylase and trypsinogen activity was 35% higher in pancreatic homogenates from
caffeine
-fed rats compared with controls. In addition, levels of immunoreactive cationic
trypsin
(ogen) were 41% higher than control levels in pancreases from the
caffeine
-fed rats. Also, the circulating levels of amylase and immunoreactive cationic
trypsin
(ogen) in serum were lower in the
caffeine
group compared with controls. When dispersed pancreatic acini isolated from the
caffeine
-fed rats were incubated in vitro with increasing concentrations of CCK-8 or nicotine, the rate of release of amylase, trypsinogen, and chymotrypsinogen was lower than in the control rats. This effect did not appear to be due to inhibition of protein synthesis, as determined by [3H]leucine incorporation into acinar protein. These data suggest that prolonged intake of
caffeine
at common dietary levels inhibits pancreatic enzyme secretion.
...
PMID:Biochemical changes in the exocrine pancreas of rats fed caffeine. 272 80
Previously, increases in ciliary length have only been obtained through genetic mutation in Chlamydomonas or by incubation of swimming echinoderm blastulae in
trypsin
or elastase. We have found that the phenotypic switch from short to long cilia on sand dollar blastulae can also be effected by incubation in theophylline. Cilia detached from control blastulae have a mean length of 21 +/- 7 microns with 10% of the cilia being greater than 30 microns. Upon incubation in 10 mM theophylline additional long cilia appeared after 10 hours and by 24-32 hours 1/2-3/4 of the embryo was covered with long cilia. The percentage of long cilia increased to 65% with a mean length of 40.0 +/- 17.6 microns. Incubation in other methylxanthines, such as aminophylline,
caffeine
, or isobutylmethylxanthine, inhibited development but had no effect on ciliary length distribution. Dibutyryl cAMP, 8-bromoadenosine, and calcium ionophore also had no effect on ciliary length. Cyclic AMP levels were measured and showed only slight differences among controls and embryos incubated in
trypsin
,
caffeine
, or theophylline. These data suggest that theophylline may be altering ciliary length control through some mechanism other than elevations in cAMP.
...
PMID:Effects of theophylline on expression of the long cilia phenotype in sand dollar blastulae. 283 65
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