| | Name | Initial Conc. (uM) | Volume (fL) | Buffered |
| 1 | CaM-GEF | 0 | 1000 | No |
| | See Farnsworth et al Nature 376 524-527 1995 |
| 2 | GAP | 0.002 | 1000 | No |
| | GTPase-activating proteins. See Boguski and McCormick. Turn off Ras by helping to hydrolyze bound GTP. This one is probably NF1, ie., Neurofibromin as it is inhibited by AA and lipids, and expressed in neural cells. p120-GAP is also a possible candidate, but is less regulated. Both may exist at similar levels. See Eccleston et al JBC 268(36) pp27012-19 Level=.002 |
| 3 | GAP* | 0 | 1000 | No |
| | |
| 4 | GDP-Ras | 0.2 | 1000 | No |
| | GDP bound form. See Rosen et al Neuron 12 1207-1221 June 1994. the activation loop is based on Boguski and McCormick Nature 366 643-654 93 Assume Ras is present at about the same level as craf-1, 0.2 uM. Hallberg et al JBC 269:6 3913-3916 1994 estimate upto 5-10% of cellular Raf is assoc with Ras. Given that only 5-10% of Ras is GTP-bound, we need similar amounts of Ras as Raf. |
| 5 | GEF* | 0 | 1000 | No |
| | phosphorylated and thereby activated form of GEF. See, e.g. Orita et al JBC 268:34 25542-25546 1993, Gulbins et al. It is not clear whether there is major specificity for tyr or ser/thr. |
| 6 | GEF-Gprot-bg | 0 | 1000 | No |
| | Guanine nucleotide exchange factor. This activates raf by exchanging bound GDP with GTP. I have left the GDP/GTP out of this reaction, it would be trivial to put them in. See Boguski & McCormick. Possible candidate molecules: RasGRF, smgGDS, Vav (in dispute). rasGRF: Kcat= 1.2/min Km = 680 nM smgGDS: Kcat: 0.37 /min, Km = 220 nM. vav: Turnover up over baseline by 10X, |
| 7 | GTP-Ras | 0 | 1000 | No |
| | Only a very small fraction (7% unstim, 15% stim) of ras is GTP-bound. Gibbs et al JBC 265(33) 20437 |
| 8 | inact-GEF | 0.1 | 1000 | No |
| | Assume that SoS is present only at 50 nM. Revised to 100 nM to get equil to experimentally known levels. |
| 9 | inact-GEF* | 0 | 1000 | No |
| | Phosphorylation-inactivated form of GEF. See Hordijk et al JBC 269:5 3534-3538 1994 and Buregering et al EMBO J 12:11 4211-4220 1993 |
