Crystal structure of M-Ras reveals a GTP-bound "off" state conformation of Ras family small GTPases.
Although some members of Ras family small GTPases, including M-Ras, share the primary structure of their effector regions with Ras, they exhibit vastly different binding properties to Ras effectors such as c-Raf-1. We have solved the crystal structure of M-Ras in the GDP-bound and guanosine 5'-(beta,gamma-imido)triphosphate (Gpp(NH)p)-bound forms. The overall structure of M-Ras resembles those of H-Ras and Rap2A, except that M-Ras-Gpp(NH)p exhibits a distinctive switch I conformation, which is caused by impaired intramolecular interactions between Thr-45 (corresponding to Thr-35 of H-Ras) of the effector region and the gamma-phosphate of Gpp(NH)p. Previous 31P NMR studies showed that H-Ras-Gpp(NH)p exists in two interconverting conformations, states 1 and 2. Whereas state 2 is a predominant form of H-Ras and corresponds to the "on" conformation found in the complex with effectors, state 1 is thought to represent the "off" conformation, whose tertiary structure remains unknown. 31P NMR analysis shows that free M-Ras-Gpp(NH)p predominantly assumes the state 1 conformation, which undergoes conformational transition to state 2 upon association with c-Raf-1. These results indicate that the solved structure of M-Ras-Gp-p(NH)p corresponds to the state 1 conformation. The predominance of state 1 in M-Ras is likely to account for its weak binding ability to the Ras effectors, suggesting the importance of the tertiary structure factor in small GTPase-effector interaction. Further, the first determination of the state 1 structure provides a molecular basis for developing novel anti-cancer drugs as compounds that hold Ras in the state 1 "off" conformation.
Division of Molecular Biology, Department of Molecular and Cellular Biology, Kobe University Graduate School of Medicine, 7-5-1 Kusunoki-cho, Chuo-ku, Kobe 650-0017, Japan.
Min
M
Ye
Fumi
F
Shima
Shin
S
Muraoka
Jingling
J
Liao
Hidetsugu
H
Okamoto
Masaki
M
Yamamoto
Atsuo
A
Tamura
Naoto
N
Yagi
Tatzuo
T
Ueki
Tohru
T
Kataoka
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D018360:Crystallography, X-Ray; D006153:Guanosine Diphosphate / Q000737:chemistry / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism*; D006165:Guanylyl Imidodiphosphate / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D020559:Monomeric GTP-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D019906:Nuclear Magnetic Resonance, Biomolecular; D017434:Protein Structure, Tertiary; D019908:Proto-Oncogene Proteins c-raf / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D020668:rap GTP-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D018631:ras Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
D006153:Guanosine Diphosphate; D006165:Guanylyl Imidodiphosphate; D006160:Guanosine Triphosphate; D019908:Proto-Oncogene Proteins c-raf; C109418:Mras protein, mouse; C492137:Rap2a protein, mouse; D020559:Monomeric GTP-Binding Proteins; D020668:rap GTP-Binding Proteins; D018631:ras Proteins