Determination of human myosin III as a motor protein having a protein kinase activity.
The class III myosin is the most divergent member of the myosin superfamily, having a domain with homology to a protein kinase. However, the function of class III myosin at a molecular level is not known at all, and it has been questioned whether it is actually an actin-based motor molecule. Here, we showed that human myosin III has an ATPase activity that is significantly activated by actin (20-fold) with Kactin of 112 microm and Vmax of 0.34 s-1, indicating the mechanoenzymatic activity of myosin III. Furthermore, we found that human myosin III has actin translocating activity (0.11 +/- 0.05 microm/s) using an in vitro actin gliding assay, and it moves toward the plus end of actin filaments. Myosin III containing calmodulin as the light chain subunit showed a protein kinase activity and underwent autophosphorylation. The autophosphorylation was the intramolecular process, and the sites were at the C-terminal end of the motor domain. Autophosphorylation significantly activated the kinase activity, although it did not affect the ATPase activity. The present study is the first report that clearly demonstrates that the class III myosin is an actin-based motor protein having a protein kinase activity.
Komaba
Shigeru
S
Department of Physiology, University of Massachusetts Medical School, Worcester, MA 01655-0127, USA.
Inoue
Akira
A
Maruta
Shinsaku
S
Hosoya
Hiroshi
H
Ikebe
Mitsuo
M
D000199:Actins / Q000378:metabolism; D000251:Adenosine Triphosphatases / Q000378:metabolism; D002850:Chromatography, Gel; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D015151:Immunoblotting; D007700:Kinetics; D025102:Myosin Type III / Q000737:chemistry* / Q000502:physiology*; D010766:Phosphorylation; D011485:Protein Binding; D011494:Protein Kinases / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D011994:Recombinant Proteins / Q000737:chemistry; D013997:Time Factors
D000199:Actins; D011994:Recombinant Proteins; D011494:Protein Kinases; D000251:Adenosine Triphosphatases; D025102:Myosin Type III