Smooth muscle myosin. Amino acid residues responsible for the hydrolysis of ATP
Onishi, H.; Morales, M.F.; Kojima, S.; Katoh, K.; Fujiwara, K.
Advances in Experimental Medicine and Biology 453: 99
1998
ISSN/ISBN: 0065-2598 PMID: 9889819 Document Number: 495835
From their crystallographic comparisons, Fisher et al. (Biochemistry 34, 8960-8972, 1995) have proposed that in an important transition of myosin heads (M), M.ATP-->M.ADP.Pi, an interdomain rotation occurs in Gly468 (of chicken smooth muscle myosin) and that the rotated state is stabilized by newly-formed interdomain contacts including the salt link between Glu470 and Arg247 (of chicken smooth muscle myosin). Here, we have studied the effects of Gly468, Glu470, and Arg247 mutations on the hydrolysis of ATP. The G468A HMM did not show a significant ATPase activity, a stoichiometric initial phosphate burst, and tryptophan fluorescence enhancement attributed to bound ADP.Pi. The E470A HMM also did not show a significant ATPase activity and the phosphate burst, but the mutant gave tryptophan response attributed to bound ATP. The E470R/R247E HMM exhibited an ATPase activity and the phosphate burst which were comparable to those of the wild-type HMM, whereas neither the E470R HMM nor the R247E HMM showed such a significant ATPase activity and burst. We thus propose that both an unhindered rotation and a salt link that stabilizes the rotated state are necessary for ATP hydrolysis.