Advertisement

Smooth Muscle Myosin

Amino Acid Residues Responsible for the Hydrolysis of ATP
  • Hirofumi Onishi
  • Manuel F. Morales
  • Shin-ichiro Kojima
  • Kazuo Katoh
  • Keigi Fujiwara
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 453)

Abstract

From their crystallographic comparisons, Fisher et al. (Biochemistry 34, 8960-8972, 1995) have proposed that in an important transition of myosin heads (M), MATP → 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.

Keywords

ATPase Activity Myosin Head Smooth Muscle Myosin E470A Mutation Essential Light Chain 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    Bagshaw, C. R. & Trentham, D. R. (1974) Biochem. J. 141, 331–349.PubMedGoogle Scholar
  2. 2.
    Bagshaw, C. R., Eccleston, J. F., Eckstein, F., Goody, R. S., Gutfreund, H., & Trentham, D. R. (1974) Biochem. J. 141, 351–364.PubMedGoogle Scholar
  3. 3.
    Fisher, A. J., Smith, C. A., Thoden, J. B., Smith, R., Sutoh, K., Holden, H. M., & Rayment, I. (1995) Biochemistry 34, 8960–8972.PubMedCrossRefGoogle Scholar
  4. 4.
    Yanagisawa, M., Hamada, Y., Katsuragawa, Y., Imamura, M., Mikawa, T. & Masaki, T. (1987).J. Mol. Biol. 198, 143–157.PubMedCrossRefGoogle Scholar
  5. 5.
    Onishi, H., Maéda, K., Maéda, Y., Inoue, A. & Fujiwara, K. (1995) Proc. Natl. Acad. Sci. USA 92, 704–708.Google Scholar
  6. 6.
    Kunkel, T. A., Roberts, J. D., & Zakour, R. A. (1987) Methods Enzymol. 154, 367–382.PubMedCrossRefGoogle Scholar
  7. 7.
    Summers, M. D. & Smith, G. E. (1987) A Manual of Methods for Baculovirus Vectors and Insect Cell Culture Procedures Bull. 1555, Texas Agric. Exp. Stn., College Station, Texas.Google Scholar
  8. 8.
    Onishi, H., Morales, M. F., Kojima, S., Katoh, K., & Fujiwara, K. (1997) Biochemistry 36, 3767–3772PubMedCrossRefGoogle Scholar
  9. 9.
    Richardson, J. S. & Richardson, D. C. (1989) in Prediction of Protein Structure and the Principles of Protein Conformation (Fasmann, G. D., ed.) pp. 1–98, Plenum Press, New York.CrossRefGoogle Scholar

Copyright information

© Plenum Press, New York 1998

Authors and Affiliations

  • Hirofumi Onishi
    • 1
  • Manuel F. Morales
    • 2
  • Shin-ichiro Kojima
    • 1
  • Kazuo Katoh
    • 1
  • Keigi Fujiwara
    • 1
  1. 1.Department of Structural AnalysisNational Cardiovascular Center Research InstituteFujishiro-dai, Suita, Osaka 565Japan
  2. 2.University of the PacificSan FranciscoUSA

Personalised recommendations