The GAP arginine finger movement into the catalytic site of Ras increases the activation entropy

@article{citeulike:2759141, abstract = {Members of the Ras superfamily of small G proteins play key roles in signal transduction pathways, which they control by GTP hydrolysis. They are regulated by GTPase activating proteins (GAPs). Mutations that prevent hydrolysis cause severe diseases including cancer. A highly conserved "arginine finger" of GAP is a key residue. Here, we monitor the GTPase reaction of the Ras{middle dot}RasGAP complex at high temporal and spatial resolution by time-resolved FTIR spectroscopy at 260 K. After triggering the reaction, we observe as the first step a movement of the switch-I region of Ras from the nonsignaling "off" to the signaling "on" state with a rate of 3 s-1. The next step is the movement of the "arginine finger" into the active site of Ras with a rate of k2 = 0.8 s-1. Once the arginine points into the binding pocket, cleavage of GTP is fast and the protein-bound Pi intermediate forms. The switch-I reversal to the "off" state, the release of Pi, and the movement of arginine back into an aqueous environment is observed simultaneously with k3 = 0.1 s-1, the rate-limiting step. Arrhenius plots for the partial reactions show that the activation energy for the cleavage reaction is lowered by favorable positive activation entropy. This seems to indicate that protein-bound structured water molecules are pushed by the "arginine finger" movement out of the binding pocket into the bulk water. The proposed mechanism shows how the high activation barrier for phosphoryl transfer can be reduced by splitting into partial reactions separated by a Pi-intermediate. 10.1073/pnas.0712095105}, author = {Kotting, Carsten and Kallenbach, Angela and Suveyzdis, Yan and Wittinghofer, Alfred and Gerwert, Klaus }, citeulike-article-id = {2759141}, doi = {10.1073/pnas.0712095105}, journal = {Proceedings of the National Academy of Sciences}, keywords = {ntpases, ras}, month = {April}, number = {17}, pages = {6260--6265}, posted-at = {2008-07-02 00:26:01}, priority = {2}, title = {The GAP arginine finger movement into the catalytic site of Ras increases the activation entropy}, url = {http://dx.doi.org/10.1073/pnas.0712095105}, volume = {105}, year = {2008} }

TY - JOUR ID - citeulike:2759141 L3 - citeulike-article-id:2759141 TI - The GAP arginine finger movement into the catalytic site of Ras increases the activation entropy JF - Proceedings of the National Academy of Sciences VL - 105 IS - 17 SP - 6260 EP - 6265 N2 - Members of the Ras superfamily of small G proteins play key roles in signal transduction pathways, which they control by GTP hydrolysis. They are regulated by GTPase activating proteins (GAPs). Mutations that prevent hydrolysis cause severe diseases including cancer. A highly conserved "arginine finger" of GAP is a key residue. Here, we monitor the GTPase reaction of the Ras{middle dot}RasGAP complex at high temporal and spatial resolution by time-resolved FTIR spectroscopy at 260 K. After triggering the reaction, we observe as the first step a movement of the switch-I region of Ras from the nonsignaling "off" to the signaling "on" state with a rate of 3 s-1. The next step is the movement of the "arginine finger" into the active site of Ras with a rate of k2 = 0.8 s-1. Once the arginine points into the binding pocket, cleavage of GTP is fast and the protein-bound Pi intermediate forms. The switch-I reversal to the "off" state, the release of Pi, and the movement of arginine back into an aqueous environment is observed simultaneously with k3 = 0.1 s-1, the rate-limiting step. Arrhenius plots for the partial reactions show that the activation energy for the cleavage reaction is lowered by favorable positive activation entropy. This seems to indicate that protein-bound structured water molecules are pushed by the "arginine finger" movement out of the binding pocket into the bulk water. The proposed mechanism shows how the high activation barrier for phosphoryl transfer can be reduced by splitting into partial reactions separated by a Pi-intermediate. 10.1073/pnas.0712095105 KW - ntpases KW - ras AU - Kotting, Carsten AU - Kallenbach, Angela AU - Suveyzdis, Yan AU - Wittinghofer, Alfred AU - Gerwert, Klaus PY - 2008/04/29/ UR - http://dx.doi.org/10.1073/pnas.0712095105 ER -