A NOVEL MECHANISM OF ACTIN ACTIVATION: ACTIN DIRECTLY ACCELERATES THE POWER-STROKE OF MYOSIN II

This thesis addresses the structural and functional aspects of the actomyosin interaction. Since the actomyosin system plays essential roles in living eukaryotic cells, the interaction between these two proteins has been of special interest for decades. In this thesis I elucidated several fundamental, but still unresolved problems in the enzymatic mechanism of the actomyosin system: Actin binding plays a fundamental role in the actomyosin ATPase cycle, because actin accelerates the rate-limiting step of the basal ATPase cycle by orders of magnitude. Therefore, the determination of the rate-limiting step is crucial for the understanding of the mechanism of actin activation. Furthermore, the assignment of the rate-limiting step will help to reveal the working mechanism of myosin and to find out whether myosins work using a Brownian-ratchet or a strain-induced conformational change mechanism. However, direct experimental evidence for the rate-limiting step in the basal ATPase cycle has been lacking. The conserved surface loop (loop 4) of myosin has been proposed but not yet experimentally demonstrated, to play a functional role in the actomyosin interaction. Our extensive kinetic and fluorescence spectroscopic analysis of loop 4 mutant myosins showed that loop 4 is a functional actin binding region of myosin and its stabilizing role is more pronounced in th