Recombinant GRF PH domain down-regulates activity of protein kinase C
- Author:
- No author available
- Journal Title:
- JOURNAL OF THE FOURTH MILITARY MEDICAL UNIVERSITY
- Issue:
- 1
- DOI:
- 10.3321/j.issn:1000-2790.2001.01.002
- Key Word:
- 信号分子;鸟苷酸类;PH结构域;蛋白激酶C;活性
Abstract： AIM To investigate association of guanine-nucleotide-releasing factor for Ras (GRF) PH domain and protein kinase C (PKC) and to examine effect of GRF PH domain on activity of protein kinase C. METHODS GRF PH domain-GST fusion protein was expressed in E.coli and purified by glutathione agarose beads. The expressed fusion protein was harvested by lysing the E.coli with ultrasonic. After centrifugation, the fusion protein in supernatant was purified by glutathione agarose beads. Western blot was employed to confirm the binding of the PH domain with PKC in vitro. To detect change of PKC activity after the fusion protein binding to PKC, PKC activity was examined with TECTTM PKC assay system. RESULTS Association of PKC with GRF PH domain was detected out by Western blot, and the PH domain down-regulated the activity of PKC. CONCLUSION Recombinant GRF PH domain binds to PKC and inhibits its kinase activity in vitro.
- [1]C A, Kanashiro,R A, Khalil.Signal transduction by protein kinase C in mammalian cells.[J].1998,25(12).
- [2]C, Shou,C L, Farnsworth,B G, Neel,etc.Molecular cloning of cDNAs encoding a guanine-nucleotide-releasing factor for Ras p21.[J].1992,358(6384).
- [3]L, Yao,Y, Kawakami,T, Kawakami.The pleckstrin homology domain of Bruton tyrosine kinase interacts with protein kinase C.[J].1994,91.
- [4]A, Toker.Signaling through protein kinase C.[J].1998,3.
- [5]R A, Mufson.The role of serine/threonine phosphorylation in hematopoietic cytokine receptor signal transduction.[J].1997,11(1).
- [6]Shibuya M,Takahashi T,Ueno H.VEGF activates protein kinase C-dependent, but Ras-independent Raf-MEK-MAP kinase pathway for DNA synthesis in primary endothelial cells.[J].1999,18(13).
- [7]Gotoh T.,Tokuda M.,Hatase O.,etc.Activation of R-Ras by Ras-Guanine Nucleotide-Releasing Factor[J].1997,272(30).
- [8]Tony Hunter.Protein kinases and phosphatases: The Yin and Yang of protein phosphorylation and signaling[J].1995,80(2).
- [9]R, Marais,Y, Light,C, Mason,etc.Requirement of Ras-GTP-Raf complexes for activation of Raf-1 by protein kinase C.[J].1998,280(5360).