期刊
AMERICAN JOURNAL OF PHYSIOLOGY-CELL PHYSIOLOGY
卷 294, 期 2, 页码 C613-C626出版社
AMER PHYSIOLOGICAL SOC
DOI: 10.1152/ajpcell.00232.2007
关键词
oxidation; proteomics
资金
- NIAMS NIH HHS [R01 AR032961, R37 AR032961, R01 AR032961-25, AR-32961] Funding Source: Medline
- NIA NIH HHS [K02 AG021626-03, R01 AG017768-05, R01 AG026160, K02 AG021626, R37 AG026160, R01 AG017768-02, AG-17768, AG-25861, AG-26160, R01 AG017768, AG-21626, K02 AG021626-05, R01 AG017768-03, K02 AG021626-02, K01 AG020990, K01 AG020990-05, R03 AG025861, R01 AG026160-04, K02 AG021626-04, R01 AG017768-04, AG-20990, R03 AG025861-02, R01 AG017768-01A1, K02 AG021626-01] Funding Source: Medline
To understand the molecular mechanism of oxidation-induced inhibition of muscle contractility, we have studied the effects of hydrogen peroxide on permeabilized rabbit psoas muscle fibers, focusing on changes in myosin purified from these fibers. Oxidation by 5 mM peroxide decreased fiber contractility (isometric force and shortening velocity) without significant changes in the enzymatic activity of myofibrils and isolated myosin. The inhibitory effects were reversed by treating fibers with dithiothreitol. Oxidation by 50 mM peroxide had a more pronounced and irreversible inhibitory effect on fiber contractility and also affected enzymatic activity of myofibrils, myosin, and actomyosin. Peroxide treatment also affected regulation of contractility, resulting in fiber activation in the absence of calcium. Electron paramagnetic resonance of spin-labeled myosin in muscle fibers showed that oxidation increased the fraction of myosin heads in the strong-binding structural state under relaxing conditions (low calcium) but had no effect under activating conditions (high calcium). This change in the distribution of structural states of myosin provides a plausible explanation for the observed changes in both contractile and regulatory functions. Mass spectroscopy analysis showed that 50 mM but not 5 mM peroxide induced oxidative modifications in both isoforms of the essential light chains and in the heavy chain of myosin subfragment 1 by targeting multiple methionine residues. We conclude that 1) inhibition of muscle fiber contractility via oxidation of myosin occurs at high but not low concentrations of peroxide and 2) the inhibitory effects of oxidation suggest a critical and previously unknown role of methionines in myosin function.
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