Access

You are not currently logged in.

Access your personal account or get JSTOR access through your library or other institution:

login

Log in to your personal account or through your institution.

If You Use a Screen Reader

This content is available through Read Online (Free) program, which relies on page scans. Since scans are not currently available to screen readers, please contact JSTOR User Support for access. We'll provide a PDF copy for your screen reader.

Identification by Mass Spectrometry of the Phosphorylated Residue Responsible for Activation of the Catalytic Domain of Myosin I Heavy Chain Kinase, a Member of the PAK/STE20 Family

Joanna Szczepanowska, Xiaolong Zhang, Christopher J. Herring, Jun Qin, Edward D. Korn and Hanna Brzeska
Proceedings of the National Academy of Sciences of the United States of America
Vol. 94, No. 16 (Aug. 5, 1997), pp. 8503-8508
Stable URL: http://www.jstor.org/stable/42886
Page Count: 6
  • Read Online (Free)
  • Subscribe ($19.50)
  • Cite this Item
Since scans are not currently available to screen readers, please contact JSTOR User Support for access. We'll provide a PDF copy for your screen reader.
Identification by Mass Spectrometry of the Phosphorylated Residue Responsible for Activation of the Catalytic Domain of Myosin I Heavy Chain Kinase, a Member of the PAK/STE20 Family
Preview not available

Abstract

Myosin I heavy chain kinase from Acanthamoeba castellanii is activated in vitro by autophosphorylation (8-10 mol of P per mol). The catalytically active C-terminal domain produced by trypsin cleavage of the phosphorylated kinase contains 2-3 mol of P per mol. However, the catalytic domain expressed in a baculovirus--insect cell system is fully active as isolated without autophosphorylation in vitro. We now show that the expressed catalytic domain is inactivated by incubation with acid phosphatase and regains activity upon autophosphorylation. The state of phosphorylation of all of the hydroxyamino acids in the catalytic domain were determined by mass spectrometry of unfractionated protease digests. Ser-627 was phosphorylated in the active, expressed catalytic domain, lost its phosphate when the protein was incubated with phosphatase, and was rephosphorylated when the dephosphorylated protein was incubated with ATP. No other residue was significantly phosphorylated in any of the three samples. Thus, phosphorylation of Ser-627, which is in the same position as the Ser and Thr residues that are phosphorylated in many other kinases, is necessary and sufficient for full activity of the catalytic domain. Ser-627 is also phosphorylated when full-length, native kinase is activated by autophosphorylation.

Page Thumbnails

  • Thumbnail: Page 
8503
    8503
  • Thumbnail: Page 
8504
    8504
  • Thumbnail: Page 
8505
    8505
  • Thumbnail: Page 
8506
    8506
  • Thumbnail: Page 
8507
    8507
  • Thumbnail: Page 
8508
    8508