Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P21817 (RyR1)
 1,154 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Analysis of the primary structure of the rabbit skeletal muscle ryanodine receptor led to the identification of two molecules of 5032 and 5037 residues, respectively. Such a sequence discrepancy is likely to be due to the alternative splicing of a 15 bp exon (1) encoding a 5 amino acid insertion (Ala-Gly-Asp-Ala-Gln) after residue 3479. By using PCR on first strand cDNA, we searched for the 15 base pair insertion in the ryanodine receptor mRNA from adult slow- and fast-twitch skeletal muscle, as well as from fast-muscles, at various stages of post-natal development. All rabbit skeletal muscle mRNAs, regardless of their developmental stage and twitch properties, contain two RYR transcripts, suggesting the coexistence of two RYR isoforms in mammalian skeletal muscle.
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PMID:Identification of two ryanodine receptor transcripts in neonatal, slow-, and fast-twitch rabbit skeletal muscles. 794 21

The potential role in Ca2+ release channel function of highly conserved, polar, and small amino acids in predicted transmembrane sequences in the rabbit skeletal muscle ryanodine receptor (RyR1) was investigated through mutagenesis. Acidic amino acids Asp3987, Glu4032, Asp4815, Asp4917, Asp4938, and Asp4969 and amidated residues Asn4034, Asn4037, Asn4574, Asn4805, Asn4806, and Gln4933, and Gly4033 were mutated to Ala, and Ala3988 was mutated to Val. When expressed in HEK-293 cells and challenged with either caffeine or 4-chloro-m-cresol, mutants E4032A, N4806A, D4815A, and D4917A did not respond, indicating that Ca2+ release channel function was impaired. None of these mutants exhibited specific binding of [3H]ryanodine. Mutants N4805A and Q4933A showed a diminished response to both caffeine and 4-chloro-m-cresol, but [3H]ryanodine binding was not altered. Other mutant responses and the responses of mutants E4032D, N4806Q or D, D4815N or E, and D4938N or E were unaltered when compared with RyR1. However, mutants E4032Q, D4917N or E, and Q4933N or E displayed neither caffeine nor 4-chloro-m-cresol response nor [3H]ryanodine binding. Sedimentation assays indicated that the nonfunctional mutants did contain tetrameric complexes, implying that defects in the assembly of a functional channel did not occur with specific mutations in transmembrane sequences. These results support the view that amino acids Glu4032 (M2), Asn4806 (M7), Asp4815 (M7), Asp4917 (M10), and Gln4933 (M10) are involved in channel function and regulation.
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PMID:Functional consequences of mutations of conserved, polar amino acids in transmembrane sequences of the Ca2+ release channel (ryanodine receptor) of rabbit skeletal muscle sarcoplasmic reticulum. 982 55

The ubiquitous glutathione transferases (GSTs) catalyze glutathione conjugation to many compounds and have other diverse functions that continue to be discovered. We noticed sequence similarities between Omega class GSTs and a nuclear chloride channel, NCC27 (CLIC1), and show here that NCC27 belongs to the GST structural family. The structural homology prompted us to investigate whether the human Omega class glutathione transferase GSTO1-1 forms or modulates ion channels. We find that GSTO1-1 modulates ryanodine receptors (RyR), which are calcium channels in the endoplasmic reticulum of various cells. Cardiac RyR2 activity was inhibited by GSTO1-1, whereas skeletal muscle RyR1 activity was potentiated. An enzymatically active conformation of GSTO1-1 was required for inhibition of RyR2, and mutation of the active site cysteine (Cys-32 --> Ala) abolished the inhibitory activity. We propose a novel role for GSTO1-1 in protecting cells containing RyR2 from apoptosis induced by Ca(2+) mobilization from intracellular stores.
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PMID:The glutathione transferase structural family includes a nuclear chloride channel and a ryanodine receptor calcium release channel modulator. 1103 31

A highly conserved amino acid sequence, GVRAGGGIGD(4831), which may form part of the Ca(2+) release channel pore in RyR2, was subjected to Ala scanning or Ala to Val mutagenesis; function was then measured by expression in HEK-293 cells, followed by Ca(2+) photometry, high affinity [(3)H]ryanodine binding, and single-channel recording. All mutants except I4829A and I4829T (corresponding to the I4897T central core disease mutant in RyR1) displayed caffeine-induced Ca(2+) release in HEK-293 cells; only mutants G4826A, I4829V, and G4830A retained high affinity [(3)H]ryanodine binding; and single-channel function was found for all mutants tested, except for G4822A and A4825V. EC(50) values for caffeine-induced Ca(2+) release were increased for G4822A, R4824A, G4826A, G4828A, and D4831A; decreased for V4823A; and unchanged for A4825V, G4827A, I4829V, and G4830A. Ryanodine (10 microm), which did not stimulate Ca(2+) release in wild type (wt), did so in Ala mutants in amino acids 4823-4827. It inhibited the caffeine response in wt and most mutants, but enhanced the amplitude of caffeine-induced Ca(2+) release in mutant G4828A. It also restored caffeine-induced Ca(2+) release in mutants I4829A and I4829T. In single-channel recordings, mutants I4829V and G4830A retained normal conductance, whereas all others had decreased unitary channel conductances ranging from 27 to 540 picosiemens. Single-channel modulation was retained in G4826A, I4829V, and G4830A, but was lost in other mutants. In contrast to wt and G4826A, I4829V, and G4830A, in which divalent metals were preferentially conducted, mutants with loss of modulation had no selectivity of divalent cations over a monovalent cation. Analysis of Gly(4822) to Asp(4831) mutants in RyR2 supports the view that this highly conserved sequence constitutes part of the ion-conducting pore of the Ca(2+) release channel and plays a key role in ryanodine and caffeine binding and activation.
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PMID:Functional characterization of mutants in the predicted pore region of the rabbit cardiac muscle Ca(2+) release channel (ryanodine receptor isoform 2). 1142 30

Although an elevation in myoplasmic Ca2+ can activate the skeletal muscle ryanodine receptor (RyR1), the function of this Ca2+ activation is unclear because extracellular Ca2+ influx is unnecessary for skeletal-type EC coupling. To determine whether Ca2+ activation of RyR1 is necessary for the initiation of skeletal-type EC coupling, we examined the behavior of RyR1 with glutamate 4032 mutated to alanine (E4032A-RyR1) because this mutation had been shown to dramatically reduce activation by Ca2+. Proc. Natl. Acad. Sci. USA. 98:2865-2870). Analysis after reconstitution into planar lipid bilayers revealed that E4032A-RyR1 was negligibly activated by 100 microM Ca2+ (P(o) too low to be measured). Even in the presence of both 2 mM caffeine and 2 mM ATP, P(o) remained low for E4032A-RyR1 (ranging from <0.0001 in 100 microM free Ca2+ to 0.005 in 2 mM free Ca2+). Thus, the E4032A mutation caused a nearly complete suppression of activation of RyR1 by Ca2+. Depolarization of E4032A-RyR1-expressing myotubes elicited L-type Ca2+ currents of approximately normal size and myoplasmic Ca2+ transients that were skeletal-type, but about fivefold smaller than those for wild-type RyR1. The reduced amplitude of the Ca2+ transient is consistent either with the possibility that Ca2+ activation amplifies Ca2+ release during EC coupling, or that the E4032A mutation generally inhibits activation of RyR1. In either case, Ca2+ activation of RyR1 does not appear to be necessary for the initiation of Ca2+ release during EC coupling in skeletal muscle.
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PMID:Ca2+ activation of RyR1 is not necessary for the initiation of skeletal-type excitation-contraction coupling. 1196 31

Phosphorylation of the skeletal muscle (RyR1) and cardiac muscle (RyR2) ryanodine receptors has been reported to modulate channel activity. Abnormally high phosphorylation levels (hyperphosphorylation) at Ser-2843 in RyR1 and Ser-2809 in RyR2 and dissociation of FK506-binding proteins from the receptors have been implicated as one of the causes of altered calcium homeostasis observed during human heart failure. Using site-directed mutagenesis, we prepared recombinant RyR1 and RyR2 mutant receptors mimicking constitutively phosphorylated and dephosphorylated channels carrying a Ser/Asp (RyR1-S2843D and RyR2-S2809D) and Ser/Ala (RyR1-S2843A and RyR2-S2809A) substitution, respectively. Following transient expression in human embryonic kidney 293 cells, the effects of Ca2+, Mg2+, and ATP on channel function were determined using single channel and [3H]ryanodine binding measurements. In both assays, neither the skeletal nor cardiac mutants showed significant differences compared with wild type. Similarly essentially identical caffeine responses were observed in Ca2+ imaging measurements. Co-immunoprecipitation and Western blot analysis showed comparable binding of FK506-binding proteins to wild type and mutant receptors. Finally metabolic labeling experiments showed that the cardiac ryanodine receptor was phosphorylated at additional sites. Taken together, the results did not support the view that phosphorylation of a single site (RyR1-Ser-2843 and RyR2-Ser-2809) substantially changes RyR1 and RyR2 channel function.
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PMID:Characterization of recombinant skeletal muscle (Ser-2843) and cardiac muscle (Ser-2809) ryanodine receptor phosphorylation mutants. 1453 76

Both imperatoxin A (IpTx(a)), a 33-residue peptide toxin from scorpion venom, and peptide A, derived from the II-III loop of dihydropyridine receptor (DHPR), interact specifically with the skeletal ryanodine receptor (RyR1), which is a Ca(2+)-release channel in the sarcoplasmic reticulum, but with considerably different affinities. IpTx(a) activates RyR1 with nanomolar affinity, whereas peptide A activates RyR1 at micromolar concentrations. To investigate the molecular basis for high-affinity activation of RyR1 by IpTx(a), we have determined the NMR solution structure of IpTx(a), and identified its functional surface by using alanine-scanning analogues. A detailed comparison of the functional surface profiles for two peptide activators revealed that IpTx(a) exhibits a large functional surface area (approx. 1900 A(2), where 1 A=0.1 nm), based on a short double-stranded antiparallel beta-sheet structure, while peptide A bears a much smaller functional surface area (approx. 800 A(2)), with the five consecutive basic residues (Arg(681), Lys(682), Arg(683), Arg(684) and Lys(685)) being clustered at the C-terminal end of the alpha-helix. The functional surface of IpTx(a) is composed of six essential residues (Leu(7), Lys(22), Arg(23), Arg(24), Arg(31) and Arg(33)) and several other important residues (His(6), Lys(8), Arg(9), Lys(11), Lys(19), Lys(20), Gly(25), Thr(26), Asn(27) and Lys(30)), indicating that amino acid residues involved in RyR1 activation make up over the half of the toxin molecule with the exception of cysteine residues. Taken together, these results suggest that the site where peptide A binds to RyR1 belongs to a subset of macrosites capable of being occupied by IpTx(a), resulting in differing the affinity and the mode of activation.
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PMID:Molecular basis of the high-affinity activation of type 1 ryanodine receptors by imperatoxin A. 1453 45

Triadin in the junctional sarcoplasmic reticulum (SR) of skeletal muscle cells has been suggested to interact with ryanodine receptor 1 (RYR1) via its KEKE motifs. Recently, we showed that amino acid residues D4878, D4907, and E4908 in RYR1 are critical for triadin-binding in vitro [J.M. Lee, S.H. Rho, D.W. Shin, C. Cho, W.J. Park, S.H. Eom, J. Ma, D.H. Kim, Negatively charged amino acids within the intraluminal loop of ryanodine receptor are involved in the interaction with triadin, J. Biol. Chem. 279 (2004) 6994-7000]. In order to test whether a disruption of the triadin-binding site(s) in RYR1 affects SR Ca(2+) release, alanine-substituted single (D4878A, D4907A, and E4908A) and triple (RYR1-TM) mutants of D4878, D4907, and E4908 were expressed in RYR1-null myotubes. Co-immunoprecipitation experiments showed a 50-60% decrease of triadin brought down in the D4907A and RYR1-TM complexes compared to the triadin-wtRYR1 complex. Ca(2+) imaging experiments using Fluo-4-AM showed atypical caffeine responses in myotubes expressing D4907A and RYR1-TM characterized by either a lack of or slower activation and faster inactivation of Ca(2+) transients. The results suggest that disruption of interaction between triadin and RYR1 impairs RYR1 function and SR Ca(2+) release.
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PMID:Occurrence of atypical Ca2+ transients in triadin-binding deficient-RYR1 mutants. 1709 84

Imperatoxin A (IpTxa), a 3.7 kDa peptide from the African scorpion Pandinus imperator, is an agonist of the skeletal muscle ryanodine receptor (RyR1). In order to study the structure of the toxin and its effect on RyR1, IpTxa cDNA was PCR-amplified using 3 pairs of primers, and the toxin was expressed in E. coli. The toxin was further purified by chromatography, and various point mutants in which basic amino acids were substituted by alanine were prepared by site-directed mutagenesis. Studies of single channel properties by the planar lipid bilayer method showed that the recombinant IpTxa was identical to the synthetic IpTxa with respect to high-performance liquid chromatography mobility, amino acid composition and specific effects on RyR1. Mutations of certain basic amino acids (Lys19, Arg23, and Arg33) dramatically reduced the capacity of the peptide to activate RyRs. A subconductance state predominated when Lys8 was substituted with alanine. These results suggest that some basic amino acid residues in IpTxa are important for activation of RyR1, and that Lys8 plays an important role in regulating the gating mode of RyR1.
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PMID:Effects of recombinant imperatoxin A (IpTxa) mutants on the rabbit ryanodine receptor. 1720 62

In the present work, we purified and characterized a novel toxin named hemicalcin from the venom of the Iranian chactoid scorpion Hemiscorpius lepturus where it represents 0.6% of the total protein content. It is a 33-mer basic peptide reticulated by three disulfide bridges, and that shares between 85 and 91% sequence identity with four other toxins, all known or supposed to be active on ryanodine-sensitive calcium channels. Hemicalcin differs from these other toxins by seven amino acids at positions 9 (leucine/arginine), 12 (alanine/glutamic acid), 13 (aspartic acid/asparagine), 14 (lysine/asparagine), 18 (serine/glycine), 26 (threonine/alanine) and 28 (proline/isoleucine/alanine). In spite of these differences, hemicalcin remains active on ryanodine-sensitive Ca2+ channels, since it increases [3H]ryanodine binding on RyR1 (ryanodine receptor type 1) and triggers Ca2+ release from sarcoplasmic vesicles. Bilayer lipid membrane experiments, in which the RyR1 channel is reconstituted and its gating properties are analysed, indicate that hemicalcin promotes an increase in the opening probability at intermediate concentration and induces a long-lasting subconductance level of 38% of the original amplitude at higher concentrations. Mice intracerebroventricular inoculation of 300 ng of hemicalcin induces neurotoxic symptoms in vivo, followed by death. Overall, these data identify a new biologically active toxin that belongs to a family of peptides active on the ryanodine-sensitive channel.
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PMID:Hemicalcin, a new toxin from the Iranian scorpion Hemiscorpius lepturus which is active on ryanodine-sensitive Ca2+ channels. 1729 Nov 97


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