Structural Dynamics of Actin during Active Interaction with Myosin depends on the Isoform of the Essential Light Chain

Abstract

Muscle contraction results from cyclic interaction of myosin and actin, driven by ATP hydrolysis, where the actomyosin complex undergoes transitions between weakly and strongly bound structural states. Rabbit skeletal muscle myosin has two isoforms of the essential light chain (ELC), designated A1 and A2. These isoforms have significant effects on myosin's interaction with actin, resulting in higher catalytic efficiency for A1 but higher in vitro motility and muscle shortening velocity for A2. In order to understand the structural basis of these differences, we have used time-resolved phosphorescence anisotropy (TPA) to investigate the effects of ELC isoforms on the microsecond dynamics of actin during interaction with skeletal muscle myosin in the absence and presence of saturating ATP. Actin was labeled with a phosphorescent probe at C374, and the myosin head (S1) was separated into isoenzymes containing A1 (S1A1) and A2 (S1A2) by ion-exchange chromatography. As previously reported, S1A1 exhibited substantially lower ATPase activity at saturating actin but substantially higher apparent actin affinity, resulting in higher catalytic efficiency. In the absence of ATP, each isoenzyme increased actin's final anisotropy similarly, indicating similar restriction of the amplitude of intrafilament rotational motions in the strong-binding (rigor) state of actomyosin. In contrast, in the presence of saturating ATP, the final anisotropy of actin was substantially higher during interaction with S1A1 than with S1A2. Thus, S1A1 was more effective in restricting actin dynamics during the active interaction of actin and myosin, suggesting greater population of the strong-binding state of actomyosin. We conclude that ELC of skeletal muscle myosin modulates strong-to-weak structural transitions during the actomyosin ATPase cycle in an isoform-dependent manner, and thus modulates the contractile function of actomyosin.