The kinetic mechanism of myole (human myosin-IC)

Myole is the widely expressed subclass-1 member of the myosin-I family. We performed a kinetic analysis of a truncated myole that consists of the motor and the single IQ motif with a bound calmodulin. We determined the rates and equilibrium constants for the key steps in the ATPase cycle. The maximum actin activated ATPase rate (V-max) and the actin concentration at half-maximum of V-max (K-ATPase) of myole are similar to those of the native protein. The K-ATPase is low (similar to1 muM), however the affinity of myole for actin in the presence of ATP is very weak. A weak actin affinity and a rapid rate of phosphate release result in a pathway under in vitro assay conditions in which phosphate is released while myole is dissociated from actin. Actin activation of the ATPase activity and the low K-ATPase are the result of actin activation of ADP release. We propose that myole is tuned to function in regions of high concentrations of cross-linked actin filaments. Additionally, we found that ADP release from actomyole is > 10-fold faster than other vertebrate myosin-I isoforms. We propose that subclass-1 myosin-Is are tuned for rapid sliding, whereas subclass-2 isoforms are tuned for tension maintenance or stress sensing.