RESUMEN
We have attempted to link the solution actomyosin ATPase with the mechanical properties of in vitro actin filament sliding over heavy meromyosin. To accomplish this we perturbed the system by altering the substrate with various NTPs and divalent cations, and by altering ionic strength. A wide variety of enzymatic and mechanical measurements were made under very similar solution conditions. Excellent correlations between the mechanical and enzymatic quantities were revealed. Analysis of these correlations based on a force-balance model led us to two fundamental equations, which can be described approximately as follows: the maximum sliding velocity is proportional to square root of V(max)K(m)(A), where K(m)(A) is the actin concentration at which the substrate turnover rate is half of its maximum (V(max)). The active force generated by a cross-bridge under no external load or under a small external load is proportional to square root of V(max)/K(m)(A). The equations successfully accounted for the correlations observed in the present study and observations in other laboratories.
Asunto(s)
Actomiosina/química , Actomiosina/metabolismo , Proteínas Motoras Moleculares/química , Proteínas Motoras Moleculares/metabolismo , Miosinas/química , Miosinas/metabolismo , Ácido Anhídrido Hidrolasas/metabolismo , Adenosina Trifosfato/metabolismo , Animales , Fenómenos Biomecánicos , Fenómenos Biofísicos , Biofisica , ATPasa de Ca(2+) y Mg(2+)/metabolismo , Cationes Bivalentes/metabolismo , Técnicas In Vitro , Cinética , Modelos Biológicos , Contracción Muscular/fisiología , Músculo Esquelético/química , Músculo Esquelético/metabolismo , Nucleósido-Trifosfatasa , Nucleótidos/metabolismo , Concentración Osmolar , Conejos , SolucionesRESUMEN
Myosin V is an unconventional myosin thought to move processively along actin filaments. To have hard evidence for the high processivity, we sought to observe directly the movement by individual native chick brain myosin V (BMV) molecules with fluorescent calmodulin. Single BMV molecules did exhibit highly processive movement along actin filaments fixed to a coverslip. BMV continued to move up to the barbed end of its actin track, and did not readily detach from action. The barbed end, therefore, got brighter with time, because of a constant stream of BMV traffic. The maximum speed of the processive movement was 1 microm/s, and the maximum actin-activated ATPase rate was 2.4 s(-1). These values apparently imply that BMV travels a great distance, 400 nm, per an ATPase cycle.