Magnesium modulates actin binding and ADP release in myosin motors

Anja M. Swenson, Darshan V. Trivedi, Anna A. Rauscher, Yuan Wang, Yasuharu Takagi, Bradley M. Palmer, András Málnási-Csizmadia, Edward P. Debold, Christopher M. Yengo

Research output: Contribution to journalArticle

15 Citations (Scopus)

Abstract

We examined the magnesium dependence of five class II myosins, including fast skeletal muscle myosin, smooth muscle myosin, β-cardiac myosin (CMIIB), Dictyostelium myosin II (DdMII), and nonmuscle myosin IIA, as well as myosin V. We found that the myosins examined are inhibited in a Mg 2+-dependent manner (0.3-9.0 mM free Mg2+) in both ATPase and motility assays, under conditions in which the ionic strength was held constant. We found that the ADP release rate constant is reduced by Mg 2+ in myosin V, smooth muscle myosin, nonmuscle myosin IIA, CMIIB, and DdMII, although the ADP affinity is fairly insensitive to Mg2+ in fast skeletal muscle myosin, CMIIB, and DdMII. Single tryptophan probes in the switch I (Trp-239) and switch II (Trp-501) region of DdMII demonstrate these conserved regions of the active site are sensitive to Mg2+ coordination. Cardiac muscle fiber mechanic studies demonstrate cross-bridge attachment time is increased at higher Mg2+ concentrations, demonstrating that the ADP release rate constant is slowed by Mg2+ in the context of an activated muscle fiber. Direct measurements of phosphate release in myosin V demonstrate that Mg2+ reduces actin affinity in the M·ADP·Pi state, although it does not change the rate of phosphate release. Therefore, the Mg2+ inhibition of the actin-Activated ATPase activity observed in class II myosins is likely the result of Mg2+- dependent alterations in actin binding. Overall, our results suggest that Mg2+ reduces the ADP release rate constant and rate of attachment to actin in both high and low duty ratio myosins.

Original languageEnglish (US)
Pages (from-to)23977-23991
Number of pages15
JournalJournal of Biological Chemistry
Volume289
Issue number34
DOIs
StatePublished - Jan 1 2014

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Myosin Type V
Myosins
Myosin Type II
Adenosine Diphosphate
Magnesium
Actins
Nonmuscle Myosin Type IIA
Skeletal Muscle Myosins
Smooth Muscle Myosins
Rate constants
Muscle
Phosphates
Switches
Cardiac Myosins
Dictyostelium
Fibers
Ionic strength
Tryptophan
Osmolar Concentration
Adenosine Triphosphatases

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Swenson, A. M., Trivedi, D. V., Rauscher, A. A., Wang, Y., Takagi, Y., Palmer, B. M., ... Yengo, C. M. (2014). Magnesium modulates actin binding and ADP release in myosin motors. Journal of Biological Chemistry, 289(34), 23977-23991. https://doi.org/10.1074/jbc.M114.562231
Swenson, Anja M. ; Trivedi, Darshan V. ; Rauscher, Anna A. ; Wang, Yuan ; Takagi, Yasuharu ; Palmer, Bradley M. ; Málnási-Csizmadia, András ; Debold, Edward P. ; Yengo, Christopher M. / Magnesium modulates actin binding and ADP release in myosin motors. In: Journal of Biological Chemistry. 2014 ; Vol. 289, No. 34. pp. 23977-23991.
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Swenson, AM, Trivedi, DV, Rauscher, AA, Wang, Y, Takagi, Y, Palmer, BM, Málnási-Csizmadia, A, Debold, EP & Yengo, CM 2014, 'Magnesium modulates actin binding and ADP release in myosin motors', Journal of Biological Chemistry, vol. 289, no. 34, pp. 23977-23991. https://doi.org/10.1074/jbc.M114.562231

Magnesium modulates actin binding and ADP release in myosin motors. / Swenson, Anja M.; Trivedi, Darshan V.; Rauscher, Anna A.; Wang, Yuan; Takagi, Yasuharu; Palmer, Bradley M.; Málnási-Csizmadia, András; Debold, Edward P.; Yengo, Christopher M.

In: Journal of Biological Chemistry, Vol. 289, No. 34, 01.01.2014, p. 23977-23991.

Research output: Contribution to journalArticle

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AU - Swenson, Anja M.

AU - Trivedi, Darshan V.

AU - Rauscher, Anna A.

AU - Wang, Yuan

AU - Takagi, Yasuharu

AU - Palmer, Bradley M.

AU - Málnási-Csizmadia, András

AU - Debold, Edward P.

AU - Yengo, Christopher M.

PY - 2014/1/1

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N2 - We examined the magnesium dependence of five class II myosins, including fast skeletal muscle myosin, smooth muscle myosin, β-cardiac myosin (CMIIB), Dictyostelium myosin II (DdMII), and nonmuscle myosin IIA, as well as myosin V. We found that the myosins examined are inhibited in a Mg 2+-dependent manner (0.3-9.0 mM free Mg2+) in both ATPase and motility assays, under conditions in which the ionic strength was held constant. We found that the ADP release rate constant is reduced by Mg 2+ in myosin V, smooth muscle myosin, nonmuscle myosin IIA, CMIIB, and DdMII, although the ADP affinity is fairly insensitive to Mg2+ in fast skeletal muscle myosin, CMIIB, and DdMII. Single tryptophan probes in the switch I (Trp-239) and switch II (Trp-501) region of DdMII demonstrate these conserved regions of the active site are sensitive to Mg2+ coordination. Cardiac muscle fiber mechanic studies demonstrate cross-bridge attachment time is increased at higher Mg2+ concentrations, demonstrating that the ADP release rate constant is slowed by Mg2+ in the context of an activated muscle fiber. Direct measurements of phosphate release in myosin V demonstrate that Mg2+ reduces actin affinity in the M·ADP·Pi state, although it does not change the rate of phosphate release. Therefore, the Mg2+ inhibition of the actin-Activated ATPase activity observed in class II myosins is likely the result of Mg2+- dependent alterations in actin binding. Overall, our results suggest that Mg2+ reduces the ADP release rate constant and rate of attachment to actin in both high and low duty ratio myosins.

AB - We examined the magnesium dependence of five class II myosins, including fast skeletal muscle myosin, smooth muscle myosin, β-cardiac myosin (CMIIB), Dictyostelium myosin II (DdMII), and nonmuscle myosin IIA, as well as myosin V. We found that the myosins examined are inhibited in a Mg 2+-dependent manner (0.3-9.0 mM free Mg2+) in both ATPase and motility assays, under conditions in which the ionic strength was held constant. We found that the ADP release rate constant is reduced by Mg 2+ in myosin V, smooth muscle myosin, nonmuscle myosin IIA, CMIIB, and DdMII, although the ADP affinity is fairly insensitive to Mg2+ in fast skeletal muscle myosin, CMIIB, and DdMII. Single tryptophan probes in the switch I (Trp-239) and switch II (Trp-501) region of DdMII demonstrate these conserved regions of the active site are sensitive to Mg2+ coordination. Cardiac muscle fiber mechanic studies demonstrate cross-bridge attachment time is increased at higher Mg2+ concentrations, demonstrating that the ADP release rate constant is slowed by Mg2+ in the context of an activated muscle fiber. Direct measurements of phosphate release in myosin V demonstrate that Mg2+ reduces actin affinity in the M·ADP·Pi state, although it does not change the rate of phosphate release. Therefore, the Mg2+ inhibition of the actin-Activated ATPase activity observed in class II myosins is likely the result of Mg2+- dependent alterations in actin binding. Overall, our results suggest that Mg2+ reduces the ADP release rate constant and rate of attachment to actin in both high and low duty ratio myosins.

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Swenson AM, Trivedi DV, Rauscher AA, Wang Y, Takagi Y, Palmer BM et al. Magnesium modulates actin binding and ADP release in myosin motors. Journal of Biological Chemistry. 2014 Jan 1;289(34):23977-23991. https://doi.org/10.1074/jbc.M114.562231