Interaction between PEVK-titin and actin filaments: origin of a viscous force component in cardiac myofibrils.

The giant muscle protein titin contains a unique sequence, the PEVK domain, the elastic properties of which contribute to the mechanical behavior of relaxed cardiomyocytes. Here, human N2-B-cardiac PEVK was expressed in Escherichia coli and tested-along with recombinant cardiac titin constructs containing immunoglobulin-like or fibronectin-like domains-for a possible interaction with actin filaments. In the actomyosin in vitro motility assay, only the PEVK construct inhibited actin filament sliding over myosin. The slowdown occurred in a concentration-dependent manner and was accompanied by an increase in the number of stationary actin filaments. High [Ca(2+)] reversed the PEVK effect. PEVK concentrations >/=10 microgram/mL caused actin bundling. Actin-PEVK association was found also in actin fluorescence binding assays without myosin at physiological ionic strength. In cosedimentation assays, PEVK-titin interacted weakly with actin at 0 degrees C, but more strongly at 30 degrees C, suggesting involvement of hydrophobic interactions. To probe the interaction in a more physiological environment, nonactivated cardiac myofibrils were stretched quickly, and force was measured during the subsequent hold period. The observed force decline could be fit with a three-order exponential-decay function, which revealed an initial rapid-decay component (time constant, 4 to 5 ms) making up 30% to 50% of the whole decay amplitude. The rapid, viscous decay component, but not the slower decay components, decreased greatly and immediately on actin extraction with Ca(2+)-independent gelsolin fragment, both at physiological sarcomere lengths and beyond actin-myosin overlap. Steady-state passive force dropped only after longer exposure to gelsolin. We conclude that interaction between PEVK-titin and actin occurs in the sarcomere and may cause viscous drag during diastolic stretch of cardiac myofibrils. The interaction could also oppose shortening during contraction.