1996 Fiscal Year Final Research Report Summary
The Significance of Posttranslational Modification (Farnesylation) of Ras Protein in Activation of Its Effectors
| Project/Area Number |
08457038
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
General medical chemistry
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| Research Institution | Kobe University School of Medicine |
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Principal Investigator |
KATAOKA Tohru Kobe Univ.Sch.Med., Dept.Physiology II,Professor, 医学部, 教授 (40144472)
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| Co-Investigator(Kenkyū-buntansha) |
KATAOKA Yuriko Kobe Univ.Sch.Med., Dept.Physiology II,Instructor, 医学部, 助手 (50233739)
KARIYA Ken-ichi Kobe Univ.Sch.Med., Dept.Physiology II,Associate Professor, 医学部, 助教授 (40263371)
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| Project Period (FY) |
1996
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| Keywords | ras Oncogene / GTP-Binding Protein / Posttranslational Modification / Rap1A / Anti-Oncogene / raf Oncogene / Adenylyl Cyclase / Adenylyl Cyclase-Associated Protein |
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| Research Abstract |
1. Based on our discovery of the second Ras-binding site of Raf-1 corresponding to the cysteine-rich region (CRR), whose interaction with Ras is abolished by mutations in the activator region of Ras and requires posttranslational modification of Ras, we elucidated the mechanism by which the anti-oncogene product Rap1A antagonizes the Ras function. Rap1A has a high affinity for CRR,forms a ternary complex with Raf-1 and Ras, and thereby inhibits the binding of Ras to CRR,resulting in inhibition of Ras-dependent Raf-1 activation. The binding of Rap1A to Raf-1 CRR also requires posttranslational modification (geranylgeranylation) of RaplA.The antagonistic function of RaplA is determined by the nature of its 31th amino acid residue, which is converted to lysine compared to glutamic acid of Ras.
2. We elucidated the molecular mechanism by which posttranslational modification (especially farnesylation) of Ras is required for activation of yeast adenylyl cyclase. Farnesylation of Ras is required for activation of adenylyl cyclase, whereas it has no effect on the binding affinity of cyclase for Ras. The stimulatory effect of farnesylation depends on the association of adenylyl cyclase with the adenylyl cyclase-associated protein CAP.This implies that CAP may be an acceptor for the farnesyl moiety of Ras and mediate the effect of Ras farnesylation. These results led us to propose a new concept of "isoprenyl group acceptor sites".
3. By employing the fluorescence polarization method, we were able to show that a synthetic peptide corresponding to the C-terminus of Ras which was chemically attached with farnesyl group bound specifically to CAP,suggesting that CAP is really an acceptor molecule for the farnesyl moiety of Ras. However, we failed to detect similar interaction of the farnesylated peptide with Raf-1 CRR.This is presumably due to vary low affinity of their interaction.
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