Divergent effects of the malignant hyperthermia-susceptible Arg(615)-->Cys mutation on the Ca(2+) and Mg(2+) dependence of the RyR1

Biophys J. 2001 Oct;81(4):2050-8. doi: 10.1016/S0006-3495(01)75854-7.

Abstract

The sarcoplasmic reticulum (SR) Ca(2+) release channel (RyR1) from malignant hyperthermia-susceptible (MHS) porcine skeletal muscle has a decreased sensitivity to inhibition by Mg(2+). This diminished Mg(2+) inhibition has been attributed to a lower Mg(2+) affinity of the inhibition (I) site. To determine whether alterations in the Ca(2+) and Mg(2+) affinity of the activation (A) site contribute to the altered Mg(2+) inhibition, we estimated the Ca(2+) and Mg(2+) affinities of the A- and I-sites of normal and MHS RyR1. Compared with normal SR, MHS SR required less Ca(2+) to half-maximally activate [(3)H]ryanodine binding (K(A,Ca): MHS = 0.17 +/- 0.01 microM; normal = 0.29 +/- 0.02 microM) and more Ca(2+) to half-maximally inhibit ryanodine binding (K(I,Ca): MHS = 519.3 +/- 48.7 microM; normal = 293.3 +/- 24.2 microM). The apparent Mg(2+) affinity constants of the MHS RyR1 A- and I-sites were approximately twice those of the A- and I-sites of the normal RyR1 (K(A,Mg): MHS = 44.36 +/- 4.54 microM; normal = 21.59 +/- 1.66 microM; K(I,Mg): MHS = 660.8 +/- 53.0 microM; normal = 299.2 +/- 24.5 microM). Thus, the reduced Mg(2+) inhibition of the MHS RyR1 compared with the normal RyR1 is due to both an enhanced selectivity of the MHS RyR1 A-site for Ca(2+) over Mg(2+) and a reduced Mg(2+) affinity of the I-site.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Binding Sites / drug effects
  • Binding Sites / physiology
  • Caffeine / pharmacology
  • Calcium / metabolism*
  • In Vitro Techniques
  • Ion Channel Gating / physiology
  • Kinetics
  • Magnesium / metabolism*
  • Malignant Hyperthermia / genetics*
  • Malignant Hyperthermia / metabolism*
  • Muscles / metabolism*
  • Point Mutation*
  • Ryanodine / metabolism
  • Ryanodine Receptor Calcium Release Channel / metabolism*
  • Swine

Substances

  • Ryanodine Receptor Calcium Release Channel
  • Ryanodine
  • Caffeine
  • Magnesium
  • Calcium