Phosphorylation-Induced Conformational Switching of CPI-17 Produces a Potent Myosin Phosphatase Inhibitor

• Published in: Structure (London) 2007-12, Vol.15 (12), p.1591-1602
• Main Authors: Eto, Masumi, Kitazawa, Toshio, Matsuzawa, Fumiko, Aikawa, Sei-ichi, Kirkbride, Jason A., Isozumi, Noriyoshi, Nishimura, Yumi, Brautigan, David L., Ohki, Shin-ya
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Animals; Circular Dichroism; Enzyme Inhibitors - chemistry; Enzyme Inhibitors - metabolism; Molecular Sequence Data; Myosin-Light-Chain Phosphatase - antagonists & inhibitors; Nuclear Magnetic Resonance, Biomolecular; Phosphorylation; Protein Conformation; PROTEINS; Sequence Homology, Amino Acid; SIGNALING; Swine
• ISSN: 0969-2126; 1878-4186
• DOI: 10.1016/j.str.2007.10.014

Phosphorylation of endogenous inhibitor proteins for type-1 Ser/Thr phosphatase (PP1) provides a mechanism for reciprocal coordination of kinase and phosphatase activities. A myosin phosphatase inhibitor protein CPI-17 is phosphorylated at Thr38 through G-protein-mediated signals, resulting in a >1000-fold increase in inhibitory potency. We show here the solution NMR structure of phospho-T38-CPI-17 with rmsd of 0.36 ± 0.06 Å for the backbone secondary structure, which reveals how phosphorylation triggers a conformational change and exposes an inhibitory surface. This active conformation is stabilized by the formation of a hydrophobic core of intercalated side chains, which is not formed in a phospho-mimetic D38 form of CPI-17. Thus, the profound increase in potency of CPI-17 arises from phosphorylation, conformational change, and hydrophobic stabilization of a rigid structure that poses the phosphorylated residue on the protein surface and restricts its hydrolysis by myosin phosphatase. Our results provide structural insights into transduction of kinase signals by PP1 inhibitor proteins.