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001-es BibID:	BIBFORM049379
Első szerző:	Lukács Balázs (élettanász)
Cím:	Effect of maurocalcine on the skeletal type ryanodine receptor / Lukacs B., Sarkozi S., Almassy J., Jona I.
Dátum:	2012
Megjegyzések:	Maurocalcine (MCa) is a 33 amino acid long scorpion toxin which shows high homology with dihydropyridine receptor constituent peptide-A (pepA). The latter is believed to directly interact with ryanodine receptor (RyR1) and plays important role in the electromechanical coupling. The position and the positive charge of given amino acids residues of MCa are determinative in this interaction.We studied the effect of pepA and potassium ion on the MCa evoked Long Lasting Subconductance State (LLSS) type RyR1 operation assuming that both of them may have access to MCa binding sites of the channel. To investigate the binding of these peptides to RyR1, we used heavy sarcoplasmic reticulum vesicles (HSR) and CHAPS solubilized ryanodine receptor complex of rabbit skeletal muscle. Gating of RyR1 was monitored on channels incorporated into a planar lipid bilayer under voltage clamp conditions. Ca2+-release measurements were performed on HSR, where changes in extravesicular Ca2+ concentration were followed as changes in the absorption of APIII Ca2+-sensitive dye (?=710 nm).In single channel experiments LLSS type RyR1 gating was evoked applying of 5 and 10 nM MCa in the cytoplasmic side of the channel. The length and frequency of the characteristic subconductance states gradually ceased by consecutively added K+. The half effective concentration of K+ was higher at 10 nM MCa concentration which refers to a possible competition between MCa and potassium ion in biding to the same site on RyR1. A similar competitive-like effect of pepA was observed, when in the presence of 26 ?M pepA, much higher concentration of MCa was able to evoke LLSSs. In Ca2+ release measurements 5 nM MCa induced Ca2+-release at 100 mM K+, but release was completely eliminated at 250 mM K+. High concentration of K+inhibited only the MCa induced Ca2+-release but had no effect on the 4-CMC induced Ca2+-release suggesting specific effect of K+ on MCa-RyR1 interaction. Suppression of release in the presence of 250 mM K+ was inhibited by addition of higher concentration of MCa suggesting charge driven interaction between MCa and RyR1.Our data put forward a possible mode of MCa action with 3 binding sites at the cytosolic side on RyR1. The first binding site located on the surface of the channel, and is responsible for the Po increase at low MCa concentration. The second binding site in the pore of the channel induces potential- and voltage dependent LLSS-s at higher toxin concentration. Occupy of third one which located presumably in the pore, close to the selectivity filter results in closed states of RyR1.Supported by: OTKA 81923
Tárgyszavak:	Természettudományok Biológiai tudományok idézhető absztrakt calcium release
Megjelenés:	Acta Physiologica. - 205 : Suppl. 690 (2012), 26. p. -
További szerzők:	Sárközi Sándor (1966-) (élettanász) Almássy János (1981-) (élettanász, biológus, angol-magyar szakfordító) Jóna István (1948-) (élettanász, fizikus)
Pályázati támogatás:	81923 OTKA
Internet cím:	Szerző által megadott URL Intézményi repozitóriumban (DEA) tárolt változat
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001-es BibID:	BIBFORM049380
Első szerző:	Sárközi Sándor (élettanász)
Cím:	Altered modulation of the skeletal type ryanodine receptor / calcium release channel by ATP in malignant hyperthermia / Sarkozi S., Lukacs B., Bardi M., Jona I.
Dátum:	2012
Megjegyzések:	We have shown previously that the activity of the Ryanodine Receptor / Calcium release channel (RyR/CRC) is modulated by ATP. Increasing ATP concentration results in an increase of open probability of the RyR/CRC and the dose response curve of the phenomena is biphasic having two distinctive activatory processes. The amplitude ratio of the two steps are 1:3 suggesting strong cooperation between the ATP binding sites and independent but cooperative binding of the ATP on the RyR1 monomers. It was also shown by several laboratories that mutations leading to malignant hyperthermia (MH) also increases the calcium sensitivity of the channel. Our aim was to test whether the ATP pharmacology is affected in MH and if this is the case in what way.Heavy SR vesicle was prepared as described previously using longissimus dorsi of a swine (Pietrin strain), which carries an MH causing homozygous arg615cysmutation. Following CHAPS+lipid solubilization, the functional RyR1 tetramer - the channel complex - was incorporated into a lipid bilayer. The bathing medium contained symmetrical 250 mM KCl ? 20mM PIPES ? pH:7.4. Free (ionized) calcium concentration was established using Ca-EGTA calcium buffer, calculated by Fabiato's method. Under voltage clamp conditions the channel current was recorded and the channel parameters were determined: such as open probability (Po), mean open time and specific conductance. The ATP pharmacology of the RyR/CRC was determined using 50 ?M Ca2+ free trans and 472 nM Ca2+ free cis, applying increasing Na2ATP concentration on the cis side.The mutant channel showed higher open probability compared to the wild type even in the absence of ATP. The mean open time was slightly higher in the mutant, but not significantly different from the wt. The ATP pharmacology of the mutant channel was different from the wt: the pronounced two phases disappeared from the ATP dependence of the open probability function. The increase of the open probability has two components: the mean open time increased significantly above 100 ?M, and the number of open events increased even more pronounced above 150 ?M ATP. The majority of the Po increase was attributed to the increase of the number of open events. All point histograms showed clearly two peaks without a trace of subconductance state: meaning that the synchrony of the four RyR1 monomers has not been changed due to the given mutation.Supported by: OTKA 81923
Tárgyszavak:	Természettudományok Biológiai tudományok idézhető absztrakt calcium release
Megjelenés:	Acta Physiologica. - 205 : Suppl. 690 (2012), p. L9. -
További szerzők:	Lukács Balázs (1978-) (élettanász) Bárdi Miklós Jóna István (1948-) (élettanász, fizikus)
Internet cím:	Szerző által megadott URL Intézményi repozitóriumban (DEA) tárolt változat
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