Mechanochemical properties of human myosin 1C are modulated by isoform-specific differences in the N-terminal extension.
Myosin-1C is a single-headed, short-tailed member of the myosin class I subfamily that supports a variety of actin-based functions in the cytosol and nucleus. In vertebrates, alternative splicing of the MYO1C gene leads to the production of three isoforms, myosin-1C0, myosin-1C16 and myosin-1C35, that carry N-terminal extensions of different length. However, it is not clear how these extensions affect the chemomechanical coupling of human myosin-1C isoforms. Here, we report on the motor activity of the different myosin-1C isoforms measuring the unloaded velocities of constructs lacking the C-terminal lipid binding domain on nitrocellulose-coated glass surfaces and full-length constructs on reconstituted, supported lipid bilayers. The higher yields of purified protein obtained with constructs lacking the lipid binding domain allowed a detailed characterization of the individual kinetic steps of human myosin-1C isoforms in their productive interaction with nucleotides and filamentous actin. Isoform-specific differences include 18-fold changes in the maximum power output per myosin-1C motor and 4-fold changes in the velocity and the resistive force at which maximum power output occurs. Our results support a model in which the isoform-specific N-terminal extensions affect chemomechanical coupling by combined steric and allosteric effects, thereby reducing both the length of the working stroke and the rate of ADP release in the absence of external loads by a factor of two for myosin-1C35 As the large change in maximum power output shows, the functional differences between the isoforms are further amplified by the presence of external loads.