Oxidation of the skeletal muscle Ca<sup>2+</sup> release  channel alters calmodulin binding

Zhang, Jia Zheng; Wu, Yifan; Williams, Barbaea Y.; Rodney, George G.; Mandel, Frederic; Strasburg, Gale M.; Hamilton, Susan L.

doi:10.1152/ajpcell.1999.276.1.c46

Cited by 74 publications

(59 citation statements)

References 39 publications

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“…The molecular mechanism of activation and inhibition of RYR1 by CaM is unknown. We have previously shown with RYR1 (10,11,18,19) CaM and apoCaM using different determinants within the same region. We have also previously shown that this region is at a subunit-subunit interface within the RYR1 tetramer (20), suggesting the possibility that CaM may regulate RYR1 function by regulating the interactions between subunits.…”

Section: Discussionmentioning

confidence: 94%

Calcium Binding to Calmodulin Leads to an N-terminal Shift in Its Binding Site on the Ryanodine Receptor

Rodney

Moore

Williams

et al. 2001

Journal of Biological Chemistry

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117

148

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The skeletal muscle calcium release channel, ryanodine receptor, is activated by calcium-free calmodulin and inhibited by calcium-bound calmodulin. Previous biochemical studies from our laboratory have shown that calcium-free calmodulin and calcium bound calmodulin protect sites at amino acids 3630 and 3637 from trypsin cleavage (Moore, C. P., Rodney, G., Zhang, J. Z., Santacruz-Toloza, L., Strasburg, G., and Hamilton, S. L. (1999) Biochemistry 38, 8532-8537). We now demonstrate that both calcium-free calmodulin and calcium-bound calmodulin bind with nanomolar affinity to a synthetic peptide matching amino acids 3614 -3643 of the ryanodine receptor. Deletion of the last nine amino acids (3635-3643) destroys the ability of the peptide to bind calcium-free calmodulin, but not calcium-bound calmodulin. We propose a novel mechanism for calmodulin's interaction with a target protein. Our data suggest that the binding sites for calcium-free calmodulin and calcium-bound calmodulin are overlapping and, when calcium binds to calmodulin, the calmodulin molecule shifts to a more N-terminal location on the ryanodine receptor converting it from an activator to an inhibitor of the channel. This region of the ryanodine receptor has previously been identified as a site of intersubunit contact, suggesting the possibility that calmodulin regulates ryanodine receptor activity by regulating subunit-subunit interactions.

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Section: Discussionmentioning

confidence: 94%

Calcium Binding to Calmodulin Leads to an N-terminal Shift in Its Binding Site on the Ryanodine Receptor

Rodney

Moore

Williams

et al. 2001

Journal of Biological Chemistry

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117

148

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“…Intersubunit Cross-linking from NEM Alkylation-The activity of RYR1 is modulated by modification of its cysteine residues by oxidation (8 -10), alkylation (11,12,19) or nitrosylation (6,7,11,20). The cysteine residues involved in these reactions have not previously been identified.…”

Section: Cam Protects Both Cysteines Required For Oxidation-inducedmentioning

confidence: 99%

“…Other sulfhydryls also react rapidly with this reagent, in particular, the cysteines that alter the ability of RYR1 to bind CaM. Alkylation of less than 5% of the total RYR1 sulfhydryls completely blocks 35 S-apoCaM binding to RYR1, without altering the ability of RYR1 to bind either 35 S-Ca 2ϩ -CaM or [ 3 H]ryanodine (12). However, both Ca 2ϩ -CaM and apoCaM can protect this site from NEM inactivation (data not shown), suggesting a steric inhibition of 35 S-apoCaM, but not 35 S-Ca 2ϩ -CaM binding.…”

Section: Cam Protects Both Cysteines Required For Oxidation-inducedmentioning

confidence: 99%

“…One of the proteins involved in excitation-contraction coupling, the skeletal muscle Ca 2ϩ -release channel, is modulated by both oxidants (8 -10) and NO (6,11). This regulation appears to be controlled, at least in part, by the binding of calmodulin (CaM) to RYR1 (12,13). Ca 2ϩ -free CaM is a partial agonist of RYR1, whereas Ca 2ϩ -CaM is an inhibitor (14).…”

mentioning

confidence: 99%

“…Ca 2ϩ -free CaM is a partial agonist of RYR1, whereas Ca 2ϩ -CaM is an inhibitor (14). In addition to these direct functional effects, CaM bound to RYR1 protects the channel from oxidation-induced intersubunit cross-linking (12), and conversely, oxidation can block CaM binding to RYR1. In contrast to the effects of oxidation, alkylation of RYR1 with NEM rapidly destroys the ability of RYR1 to bind 35 S-Ca 2ϩ -free CaM (apoCaM) but does not alter its ability to bind 35 S-Ca 2ϩ -CaM.…”

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confidence: 99%

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A Role for Cysteine 3635 of RYR1 in Redox Modulation and Calmodulin Binding

Moore

Zhang

Hamilton

1999

Journal of Biological Chemistry

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In the current work, we demonstrate that both cysteines needed for the oxidation-induced intersubunit crosslink are protected from alkylation with N-ethylmaleimide by bound calmodulin. We also show, using N-terminal amino acid sequencing together with analysis of the distribution of [ 3 H]NEM labeling with each sequencing cycle, that cysteine 3635 of RYR1 is rapidly labeled by NEM and that this labeling is blocked by bound calmodulin. We propose that cysteine 3635 is located at an intersubunit contact site that is close to or within a calmodulin binding site. These findings suggest that calmodulin and oxidation modulate RYR1 activity by regulating intersubunit interactions in a mutually exclusive manner and that these interactions involve cysteine 3635.Reactive oxygen species and nitric oxide (NO) 1 are produced by skeletal muscle even at rest, but their levels are dramatically increased by muscle activity (1-5). Both reactive oxygen species and NO alter muscle function (6), possibly by altering excitation-contraction coupling (7). One of the proteins involved in excitation-contraction coupling, the skeletal muscle Ca 2ϩ -release channel, is modulated by both oxidants (8 -10) and NO (6,11). This regulation appears to be controlled, at least in part, by the binding of calmodulin (CaM) to RYR1 (12, 13). Ca 2ϩ -free CaM is a partial agonist of RYR1, whereas Ca 2ϩ -CaM is an inhibitor (14). In addition to these direct functional effects, CaM bound to RYR1 protects the channel from oxidation-induced intersubunit cross-linking (12), and conversely, oxidation can block CaM binding to RYR1. In contrast to the effects of oxidation, alkylation of RYR1 with NEM rapidly destroys the ability of RYR1 to bind 35 S-Ca 2ϩ -free CaM (apoCaM) but does not alter its ability to bind 35 S-Ca 2ϩ -CaM. Alkylation also prevents oxidation-induced intersubunit crosslinking. Our studies suggest that there is only one CaM site per subunit of RYR1 at either high or low Ca 2ϩ (13). Some indication of the location of the CaM binding site has been obtained from examination of sites on RYR1 protected from tryptic cleavage by CaM (13). Treatment of RYR1 with trypsin rapidly destroys its ability to bind CaM, but CaM bound to RYR1 can protect its binding site from tryptic digestion. The sites protected by Ca 2ϩ -CaM or apoCaM are at amino acids 3630 and 3637, suggesting that either the binding sites for CaM in both the Ca 2ϩ -free and Ca 2ϩ -bound forms are physically close to this region of RYR1 or that the binding of both forms of CaM produces a conformational change that buries this region of RYR1. The latter possibility seems unlikely because the functional effects of apoCaM are opposite those of Ca 2ϩ -CaM. The sequence between these two sites is AVVACFR, suggesting that cysteine 3635 may be one of the cysteines that, in the absence of CaM, can form the intersubunit disulfide bond. This intersubunit contact site is likely to represent an important site for regulating RYR1 activity. In the current study, we demonstrate that cysteine 3635 is one...

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Reactive Oxygen Species: Impact on Skeletal Muscle

Powers

Kavazis

et al. 2011

Comprehensive Physiology

392

384

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It is well established that contracting muscles produce both reactive oxygen and nitrogen species. Although the sources of oxidant production during exercise continue to be debated, growing evidence suggests that mitochondria are not the dominant source. Regardless of the sources of oxidants in contracting muscles, intense and prolonged exercise can result in oxidative damage to both proteins and lipids in the contracting myocytes. Further, oxidants regulate numerous cell signaling pathways and modulate the expression of many genes. This oxidant-mediated change in gene expression involves changes at transcriptional, mRNA stability, and signal transduction levels. Furthermore, numerous products associated with oxidant-modulated genes have been identified and include antioxidant enzymes, stress proteins, and mitochondrial electron transport proteins. Interestingly, low and physiological levels of reactive oxygen species are required for normal force production in skeletal muscle, but high levels of reactive oxygen species result in contractile dysfunction and fatigue. Ongoing research continues to explore the redox-sensitive targets in muscle that are responsible for both redox-regulation of muscle adaptation and oxidant-mediated muscle fatigue.

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Oxidation of the skeletal muscle Ca²⁺ release channel alters calmodulin binding

Cited by 74 publications

References 39 publications

Calcium Binding to Calmodulin Leads to an N-terminal Shift in Its Binding Site on the Ryanodine Receptor

Calcium Binding to Calmodulin Leads to an N-terminal Shift in Its Binding Site on the Ryanodine Receptor

A Role for Cysteine 3635 of RYR1 in Redox Modulation and Calmodulin Binding

Reactive Oxygen Species: Impact on Skeletal Muscle

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Oxidation of the skeletal muscle Ca2+ release channel alters calmodulin binding

Cited by 74 publications

References 39 publications

Calcium Binding to Calmodulin Leads to an N-terminal Shift in Its Binding Site on the Ryanodine Receptor

Calcium Binding to Calmodulin Leads to an N-terminal Shift in Its Binding Site on the Ryanodine Receptor

A Role for Cysteine 3635 of RYR1 in Redox Modulation and Calmodulin Binding

Reactive Oxygen Species: Impact on Skeletal Muscle

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Product

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Oxidation of the skeletal muscle Ca²⁺ release channel alters calmodulin binding