Hiroyuki Matsumiya

The effect of EDTA on the interaction between actomyosin and adenosine triphosphate

Abstract 1. 1. In the presence of 0.6M of KCl, EDTA increases concomitantly the Vmax and Km of myosin B ATPase. Especially when concentration of EDTA is higher than 5mM, the line 1/v versus 1/[S] passes through the origin. 2. 2. EDTA decreases both grade and rate of the optical change of myosin B caused by ATP addition. 3. 3. The recovery step from the physically changed myosin B to the original myosin B follows strictly monomolecular kinetics. 4. 4. On the basis of these results the reaction mechanism of EDTA with the myosin B-ATP system is discussed.

Enzymatic deamination of adenosine 2′,3′-cyclic phosphate in Drosophilamelanogaster

Abstract Four nucleoside cyclic phosphates were isolated from the hot ethanol extracts of the third instar larvae of Drosophila melanogaster and identified as follows: cytidine 2′,3′-cyclic phosphate(Cp!), uridine 2′,3′-cyclic phosphate(Up!), guanosine 2′,3′-cyclic phosphate(Gp!) and inosine 2′,3′-cyclic phosphate(Ip!). The occurrence of Ip! instead of Ap! in the larvae suggests the presence of a deaminase which catalyzes the conversion of Ap! to Ip!. Such an enzyme has indeed been shown to be present in Drosophila larvae.

Rotary diffusion of actomyosin

Abstract 1. 1. The viscosity, the intensity of flow birefringence, and the rotary diffusion constant of myosin B, prepared from rabbit skeletal muscle, were investigated over a wide range of the velocity gradient (17–12,500 sec−1). The following results were obtained. 2. 2. The viscosity decreased with increase of the gradient. 3. 3. At low gradient myosin B particles behaved polydispersedly with respect to the rotary diffusion, while at high gradient they were monodisperse. The rotary diffusion constant increased with increment of the gradient, and at sufficiently high gradient it was a constant value (ca. 20 sec−1), independent of gradient and the protein concentration. 4. 4. The length of the equivalent elongated ellipsoid calculated from the rotary diffusion constant at sufficiently high gradient was 8,600–10,100 A, in satisfactory agreement with results obtained from light scattering. 5. 5. At a low range of gradient ( 6. 6. If the protein solution were once subjected to a high gradient, the intensity of flow birefringence observed at low gradient was considerably higher than the original value. 7. 7. In high KCl solution, the rotary diffusion constant was observed to be higher than 10 sec−1 even at a low range of gradient. 8. 8. Consequently, it was concluded that a network is built up by the electrostatic interferences of the constituent elementary particles with each other and the entanglements are broken more and more as the gradient increases, that the length of the elementary particles is almost constant, and that the orientation of polypeptide chains in these particles is improved by high shearing stress. 9. 9. The rotary diffusion constant at high shearing stress increased remarkably on the addition of ATP. The molecular mechanism of this effect was briefly discussed.

The reaction of phenylmethanesulfonyl-subtilisin

Abstract 1. 1.|Phenylmethanesulfonyl fluoride-inactivated subtilisin is reactivated by formohydroxamic acid and hydroxylamine. Formohydroxamic acid causes reactivation to an extent of 100%. 2. 2.|From the observation that the pH-activity profile of reactivation of the inactivated subtilisin by formohydroxamic acid was S-shaped, it is concluded that the reactivation is analogous to the deacylation step of the enzyme-substrate reaction. 3. 3.|Diisopropylfluorophosphate-inactivated subtilisin is not reactivated by formohydroxamic acid.