Peroxynitrite inhibits myofibrillar protein function in an in vitro assay of motility.
We determined the effects of peroxynitrite (ONOO-) on cardiac myosin, actin, and thin filaments in order to more clearly understand the impact of this reactive compound in ischemia/reperfusion injury and heart failure. Actin filaments, native thin filaments, and alpha-cardiac myosin from rat hearts were exposed to ONOO- in the presence of 2 mM bicarbonate. Filament velocities over myosin, calcium sensitivity, and relative force generated by myosin were assessed in an in vitro motility assay in the absence of reducing agents. ONOO- concentrations > or =10 microM significantly reduced the velocities of thin filaments or bare actin filaments over alpha-cardiac myosin when any of these proteins were exposed individually. These functional deficits were linearly related to the degree of tyrosine nitration, with myosin being the most sensitive. However, at 10 microM ONOO- the calcium sensitivity of thin filaments remained unchanged. Cotreatment of myosin and thin filaments, analogous to the in vivo situation, resulted in a significantly greater functional deficit. The load supported by myosin after ONOO- exposure was estimated using mixtures experiments to be increased threefold. These data suggest that nitration of myofibrillar proteins can contribute to cardiac contractile dysfunction in pathologic states in which ONOO- is liberated.
Department of Biomedical Engineering, University of Virginia, Charlottesville, VA 22908, USA.
Jeremy H
JH
Snook
Jiahui
J
Li
Brian P
BP
Helmke
William H
WH
Guilford
D000199:Actins / Q000187:drug effects / Q000502:physiology; D000465:Algorithms; D000818:Animals; D002118:Calcium / Q000378:metabolism; D024722:Cardiac Myosins / Q000187:drug effects / Q000502:physiology; D066298:In Vitro Techniques; D008958:Models, Molecular; D020409:Molecular Motor Proteins / Q000187:drug effects* / Q000502:physiology; D009200:Myocardial Contraction / Q000187:drug effects / Q000502:physiology; D015428:Myocardial Reperfusion Injury / Q000378:metabolism / Q000503:physiopathology; D009210:Myofibrils / Q000187:drug effects* / Q000378:metabolism; D018384:Oxidative Stress; D030421:Peroxynitrous Acid / Q000378:metabolism / Q000494:pharmacology*; D051381:Rats; D017382:Reactive Oxygen Species
D000199:Actins; D020409:Molecular Motor Proteins; D017382:Reactive Oxygen Species; D030421:Peroxynitrous Acid; D024722:Cardiac Myosins; D002118:Calcium