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 journalArticlepeer-review

20 Scopus citations

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 - 2014

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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