PubMed Health⌕ Search

Biomedical subjects

V Shoshan-Barmatz

Publications and source records attributed to V Shoshan-Barmatz.

48 records · Page 3Linked to original sources
PubMed1989

High-affinity Ca2+-binding site inhibiting Ca2+ release from sarcoplasmic reticulum.

Ca2+ release from sarcoplasmic reticulum membranes, activated by alkaline pH occurs only when EGTA is present in the release medium. Addition of very low concentrations of Ca2+ to the medium inhibits Ca2+ release. The concentration of free Ca2+ required for 50% inhibition ranges from between 5 and 20 nM in different experiments and/or membrane preparations, irrespective of whether the free Ca2+ concentration is controlled by EGTA or CDTA. Other divalent cations such as Mn2+, Ba2+, Cu2+, Cd2+ and Mg2+ also exert an inhibitory effect on Ca2+ release, with higher or lower potency than that of Ca2+. The inactivation of Ca2+ release by Ca2+ is reversible. We suggest the involvement of high-affinity Ca2+-binding sites in the control of Ca2+ release.

Calcium↗
PubMed1988

ATP-dependent interaction of propranolol and local anaesthetic with sarcoplasmic reticulum. Stimulation of Ca2+ efflux.

Preincubation of sarcoplasmic reticulum (SR) with propranolol or tetracaine inhibits Ca2+ accumulation and stimulates ATPase activity by more than 2-fold. This effect is obtained only when the preincubation is carried out in the presence of ATP or other nucleoside triphosphates. The (ATP + drug)-induced inhibition of Ca2+ accumulation is pH-dependent, increasing as the pH rises above 7.5. The presence of micromolar concentrations of Ca2+ or Mg2+ during the preincubation prevents the inhibitory effect of ATP plus drug on Ca2+ accumulation or ATPase activity. The (ATP + drug) modification of SR vesicles resulted in stimulation of a rapid Ca2+ efflux from passively loaded vesicles. The ATP-dependent inhibition of Ca2+ accumulation by the drug is obtained with other local anaesthetics. The drug concentration required for 50% inhibition was 0.15 mM for dibucaine and 0.4 mM for both propranolol and tetracaine, whereas it was 5 mM, 8 mM and greater than 10 mM for lidocaine, benzocaine and procaine respectively. The heavy SR vesicles were only slightly affected by the incubation with propranolol or tetracaine in the presence of ATP, but their sensitivity increased markedly after storage at 0 degrees C for 24-48 h. These results suggest that propranolol and some local anaesthetics, in the presence of ATP, stimulate Ca2+ efflux by modifying a protein factor(s) rather than the phospholipid bilayer.

Adenosine Triphosphatases↗
PubMed1988

Trypsin destruction of the high affinity ryanodine binding sites of the junctional sarcoplasmic reticulum.

Tryptic digestion of the junctional sarcoplasmic reticulum membranes in sucrose but not NaCl buffer leads to complete loss of ryanodine binding capacity. The presence of MgCl2 in the sucrose buffer prevents the loss of ryanodine binding by the trypsin treatment. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the treated membranes reveal that the 400-kDa protein band disappeared under all the different digestion conditions. However, the presence of 135-kDa tryptic fragment is observed only when ryanodine binding is retained. Quantitative analysis of the gels shows that the loss of ryanodine binding is well correlated with the cleavage of the 135-kDa tryptic fragment. This correlation is obtained when the cleavage was controlled either by the digestion time or by NaCl or MgCl2 concentrations. The same concentrations of MgCl2 and NaCl affect the ryanodine binding activity, the cleavage of the 135-kDa tryptic fragment, and the solubility and stability of the [3H]ryanodine-receptor complex in a detergent-containing medium. Tryptic digestion of the ryanodine receptor/junctional Ca2+ release channel, which leads to complete loss of ryanodine binding capacity, has no effect or slightly stimulates the Ca2+ accumulation activity of these membranes.

Adenosine Triphosphate↗
PubMed1988

Dicyclohexylcarbodiimide interaction with sarcoplasmic reticulum. Inhibition of Ca2+ efflux.

Dicyclohexylcarbodiimide (DCCD), a hydrophobic carboxyl reagent, inhibited Ca2+ release from Ca2+-loaded sarcoplasmic reticulum vesicles, induced by elevated pH, tetraphenylboron, ATP + Pi, or membrane modification with acetic anhydride. Under the conditions used, the same concentrations of DCCD were required for inhibition of Ca2+ release, Ca2+-ATPase activity, and Ca2+ uptake. On the other hand, free Ca2+ or alkaline pH prevented the inhibition by DCCD of Ca2+-ATPase and coupled Ca2+ transport but not that of Ca2+ release. Moreover, several hydrophilic carboxyl reagents inhibited Ca2+-ATPase but not Ca2+ release. We suggest that a carboxyl residue(s), located in a hydrophobic region of a protein(s), is involved in the control of Ca2+ release, where DCCD interaction with this group blocks Ca2+ release. This group is distinct from the one involved in the inhibition of Ca2+-ATPase. DCCD also inhibited [3H]ryanodine binding to junctional sarcoplasmic reticulum membranes. The presence of Ca2+ or an alkaline pH only slightly affects the degree of inhibition of ryanodine binding by DCCD. Incubation of the membranes with [14C]DCCD resulted in labeling of 350-, 170-, 140-, 53-, and 30-kDa proteins in addition to the Ca2+-ATPase. The involvement of one or all of the DCCD-labeled proteins in Ca2+ release and ryanodine binding is discussed.

Acetic Anhydrides↗
PubMed1988

Ryanodine binding to sarcoplasmic reticulum membrane; comparison between cardiac and skeletal muscle.

[3H]Ryanodine binding to skeletal muscle and cardiac sarcoplasmic reticulum (SR) vesicles was compared under experimental conditions known to inhibit or stimulate Ca2+ release. In the skeletal muscle SR, ryanodine binds to a single class of high-affinity sites (Kd of 11.3 nM). In cardiac SR vesicles, more than one class of binding sites is observed (Kd values of 3.6 and 28.1 nM). Ryanodine binding to skeletal muscle SR vesicles requires high concentrations of NaCl, whereas binding of the drug to cardiac SR is only slightly influenced by ionic strength. In the presence of 5'-adenylyl imidodiphosphate (p[NH]ppA), increased pH, and micromolar concentration of Ca2+ (which all induce Ca2+ release from SR) binding of ryanodine to SR is significantly increased in skeletal muscle, while being unchanged in cardiac muscle. Ryanodine binding to skeletal but not to cardiac muscle SR is inhibited in the presence of high Ca2+ or Mg2+ concentrations (all known to inhibit Ca2+ release from skeletal muscle SR). Ruthenium red or dicyclohexylcarbodiimide modification of cardiac and skeletal muscle SR inhibit Ca2+ release and ryanodine binding in both skeletal and cardiac membranes. These results indicate that significant differences exist in the properties of ryanodine binding to skeletal or cardiac muscle SR. Our data suggest that ryanodine binds preferably to site(s) which are accessible only when the Ca2+ release channel is in the open state.

Adenylyl Imidodiphosphate↗
PubMed1988

Activation of Ca2+ release in isolated sarcoplasmic reticulum.

The relationship between Ca2+ release from sarcoplasmic reticulum, induced by elevated pH, tetraphenylboron (TPB-) or chemical modification, and the change in the surface charge of the membranes as measured by the fluorescence intensity of anilinonaphthalene sulfonate (ANS) is examined. The simulated Ca2+ release is inhibited by dicyclohexylcarbodiimide and external Ca2+. TPB-, but not tetraphenylarsonium (TPA+), causes a decrease in ANS- fluorescence, with 50% decrease occurring at about 5 microM TPB-. The decrease in ANS- fluorescence as well as the inhibition of Ca2+ accumulation induced by TPB- are prevented by TPA+. A linear relationship between the decrease in membrane surface potential and the extent of the Ca2+ released by TPB- is obtained. Similar levels of [3H]TPB-bound to sarcoplasmic reticulum membranes were obtained regardless of whether or not the vesicles have taken up Ca2+. The inhibition of Ca2+ accumulation and the [3H]TPB- incorporation into the membranes were correlated. Ca2+ release from sarcoplasmic reticulum, by pH elevation, chemical modification or by addition of NaSCN (0.2 to 0.5 M) or the Ca2+ ionophore ionomycin, is also accompanied by a decrease in ANS- fluorescence intensity. However, chemical modification and elevated pH affects the surface potential much less than SCN- or TPB- do. These results suggest that the enhancement of Ca2+ release by these treatments is not due to a general effect on the membrane surface potential, but rather through the modification of a specific protein. They also suggest that membrane surface charges might play an important role in the control mechanism of Ca2+ release.

Anilino Naphthalenesulfonates↗
PubMed1987

Stimulation of Ca2+ efflux from sarcoplasmic reticulum by preincubation with ATP and inorganic phosphate.

Preincubation of sarcoplasmic reticulum with 1 mM-ATP completely inhibits Ca2+ accumulation and stimulates ATPase activity by over 2-fold. This effect of ATP is obtained only when the preincubation is carried out in the presence of Pi, but not with arsenate, chloride or sulphate. The inhibition by ATP of Ca2+ accumulation is pH-dependent, increasing as the pH is increased above 7.5. Inhibition of Ca2+ accumulation is observed on preincubation with ATP, but not with CTP, UTP, GTP, ADP, adenosine 5'-[beta gamma-methylene]triphosphate or adenosine 5'-[beta gamma-imido]triphosphate. The presence of Ca2+, but not Mg2+, during the preincubation, prevents the effect of ATP + Pi on Ca2+ accumulation. The ATP + Pi inhibition of Ca2+ accumulation is not due to modification of the ATPase catalytic cycle, but rather to stimulation of a rapid Ca2+ efflux from actively or passively loaded vesicles. This Ca2+ efflux is inhibited by dicyclohexylcarbodi-imide. Photoaffinity labelling of sarcoplasmic-reticulum membranes with 8-azido-[alpha-32P]ATP resulted in specific labelling of two proteins, of approx. 160 and 44 kDa. These proteins were labelled in the presence of Pi, but not other anions.

Adenosine Triphosphatases↗
PubMed1987

Tryptic digestion of sarcoplasmic reticulum inhibits Ca2+ accumulation by action on a membrane component other than the Ca2+-ATPase.

We have reexamined the "uncoupling" of Ca2+ transport from ATP hydrolysis, which has been reported to be caused by trypsin cleavage of the Ca2+-ATPase of sarcoplasmic reticulum (SR) vesicles at the second (slower) of two characteristic tryptic sites (Scott, T. L., and Shamoo, A. E. (1982) J. Membr. Biol. 64, 137-144). We find that the loss of Ca2+ accumulation capacity in SR vesicles is poorly correlated with this cleavage under several conditions. The loss is accompanied by increased Ca2+ permeability but not by changes in the properties of the ATPase or ATP-Pi exchange activities of the vesicles. Proteoliposomes containing purified Ca2+-ATPase which has been cleaved in part at the two tryptic sites are as well coupled and impermeable to Ca2+ as proteoliposomes containing intact Ca2+-ATPase. We conclude that the loss of Ca2+ accumulation capacity in SR vesicles on tryptic treatment is due to cleavage of a SR membrane component other than the Ca2+-ATPase, possibly a component of the gated channels which function in Ca2+ release from SR, which leads to a Ca2+ leak. The hydrolytic and coupled transport functions of the Ca2+-ATPase itself may well be unaffected by the two tryptic cleavages.

Animals↗
PubMed1987

Chemical modification of sarcoplasmic reticulum with methylbenzimidate. Stimulation of Ca2+ efflux.

Treatment of sarcoplasmic reticulum membranes with 12 mM-methylbenzimidate (MBI) for 5 min, in the presence of 5 mM-ATP at pH 8.5, resulted in a 2-3-fold stimulation of ATP hydrolysis and over 90% inhibition of Ca2+ accumulation. This phenomenon was strictly dependent upon the presence of nucleotides with the following order of effectiveness: adenosine 5'-[beta, gamma-imido]triphosphate greater than or equal to ATP greater than UTP greater than ADP greater than AMP. Divalent cations such as Ca2+, Mg2+ and Mn2+, when present during the MBI treatment, prevented both the stimulation of ATPase activity and the inhibition of Ca2+ accumulation. Modification with MBI had no effect on E-P formation from ATP, ADP-ATP exchange, Ca2+ binding or ATP-Pi exchange catalysed by the membranes. Membranes modified with MBI in the presence of ATP and then passively loaded with Ca2+ released about 80% of their Ca2+ content within 3 s. Control membranes released only 3% of their Ca2+ during the same time period. MBI modification inhibited Ca2+ accumulation by proteoliposomes reconstituted with the partially purified ATPase but not with the purified ATPase fraction. These results suggest that MBI in the presence of ATP stimulates Ca2+ release by modifying a protein factor(s) other than the (Ca2+ + Mg2+)-ATPase.

Adenosine Triphosphatases↗
PubMed1986

Chemical modification of sarcoplasmic reticulum. Stimulation of Ca2+ release.

Pretreatment of sarcoplasmic membranes with acetic or maleic anhydrides, which interact principally with amino groups, resulted in an inhibition of Ca2+ accumulation and ATPase activity. The presence of ATP, ADP or adenosine 5'-[beta, gamma-imido]triphosphate in the modification medium selectively protected against the inactivation of ATPase activity by the anhydride but did not protect against the inhibition of Ca2+ accumulation. Acetic anhydride modification in the presence of ATP appeared to increase specifically the permeability of the sarcoplasmic reticulum membrane to Ca2+ but not to sucrose, Tris, Na+ or Pi. The chemical modification stimulated a rapid release of Ca2+ from sarcoplasmic reticulum vesicles passively or actively loaded with calcium, from liposomes reconstituted with the partially purified ATPase fraction but not from those reconstituted with the purified ATPase. The inactivation of Ca2+ accumulation by acetic anhydride (in the presence of ATP) was rapid and strongly pH-dependent with an estimated pK value above 8.3 for the reactive group(s). The negatively charged reagents pyridoxal 5-phosphate and trinitrobenzene-sulphonate, which also interact with amino groups, did not stimulate Ca2+ release. Since these reagents do not penetrate the sarcoplasmic reticulum membranes, it is proposed that Ca2+ release is promoted by modification of internally located, positively charged amino group(s).

Acetic Anhydrides↗
PubMedDate not supplied

The action of carboxyl modifying reagents on the ryanodine receptor/Ca2+ release channel of skeletal muscle sarcoplasmic reticulum.

In this work we show that ryanodine binding to junctional sarcoplasmic reticulum (SR) membranes or purified ryanodine receptor (RyR) is inhibited in a time- and concentration-dependent fashion by prior treatment with the carboxyl reagent dicyclohexylcarbodiimide (DCCD). Exposure of the membrane-bound RyR to the water soluble carboxyl reagents 1-ethyl-3 (3-(dimethylamino) propyl carbodiimide (EDC) or N-ethyl-pheny-lisoxazolium-3'-sulfonate (WRK) only slightly affects their ryanodine binding capacity. The amphipathic reagent N-ethoxy cabonyl-2-ethoxy-1,2-dihydroquinaline (EEDQ) inhibited ryanodine binding at relatively high concentrations. DCCD-modification of the SR decreased the binding affinities of the RyR for ryanodine and Ca2+ by about 3- and 18-fold, respectively. The single channel activity of SR membranes modified with DCCD and then incorporated into planar lipid bilayers is very low (5-8%) in comparison to control membranes. Application of DCCD to either the myoplasmic (cis) or luminal (trans) side of the reconstituted unmodified channels resulted in complete inhibition of their single channel activities. Similar results were obtained with the water soluble reagent WRK applied to the myoplasmic, but not to the luminal side. The DCCD-modified non-active channel is re-activated by addition of ryanodine in the presence of 250 microM Ca2+ and is stabilized in a sub-conductance state. With caffeine, ryanodine re-activated the channel in the presence of 100 microM of Ca2+. The results suggest that a carboxyl residue(s) in the RyR is involved either in the binding of Ca2+, or in conformational changes that are produced by Ca2+ binding, and are required for the binding of ryanodine and the opening of the Ca2+ release channel.

Animals↗