Parallel coiled-coil association of the RhoA-binding domain in Rho-kinase.
Rho-kinase is a serine/threonine protein kinase that regulates cytoskeletal events in cells. The enzyme activity of Rho-kinase is auto-inhibited in the free state but is activated through direct binding to the small GTPase Rho in the GTP-bound form. The crystal structure of the Rho-binding domain (RhoBD) of Rho-kinase has been determined at 1.8-A resolution by the multi-wavelength anomalous dispersion technique. The structure shows that RhoBD dimerizes to form a parallel coiled-coil with long consecutive alpha-helices extended to approximately 97 A and suggests that free Rho-kinase can also form a dimer through parallel self-association. At the middle region of the coiled-coil, the polypeptide chains are flexible and display loose "knobs-into-holes" packing of the side chains from both chains. RhoBD residues that have been shown to be critical for Rho-binding are spread in the positively charged C-terminal region. The parallel coiled-coil structure of our Rho-kinase RhoBD in the free form is different from the anti-parallel coiled-coil structure of RhoBD of protein kinase N when complexed with RhoA. Implications derived from these structural studies in relation to the mechanism of Rho-kinase activation will be addressed with previously reported experimental data.
Shimizu
Toshiyuki
T
Structural Biology Laboratory, Nara Institute of Science and Technology, and CREST, Japan Science and Technology Corporation, 8916-5 Takayama, Ikoma, Nara 630-0192, Japan.
Ihara
Kentaro
K
Maesaki
Ryoko
R
Amano
Mutsuki
M
Kaibuchi
Kozo
K
Hakoshima
Toshio
T
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D018360:Crystallography, X-Ray; D003599:Cytoskeleton / Q000378:metabolism; D019281:Dimerization; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D011493:Protein Kinase C / Q000737:chemistry; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry*; D017434:Protein Structure, Tertiary; D051381:Rats; D054460:rho-Associated Kinases; D020742:rhoA GTP-Binding Protein / Q000737:chemistry* / Q000502:physiology
D047908:Intracellular Signaling Peptides and Proteins; D010455:Peptides; D016244:Guanosine 5'-O-(3-Thiotriphosphate); C061570:protein kinase N; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D011493:Protein Kinase C; D020742:rhoA GTP-Binding Protein