Equilibrium and Kinetic Study of the Conformational Transition toward the Active State of p21Ha-, Induced by the Binding of BeF3− to the GDP-bound State, in the Absence of GTPase-activating Proteins

Abstract

). The similarity between the GTPbound form and the GDP⅐BeF 3؊ -bound form has been confirmed using lifetime analysis of the tryptophan fluorescence. The kinetic analysis of the process indicates that the binding can be divided into a first bimolecular step, which accounts for the association of the anion with its binding site, and a second step, which corresponds to an internal conformational transition of the GDP⅐BeF 3 ؊ ⅐p21 Ha-ras complex to its final state. Both steps are endothermic (⌬H 1 ‫؍‬ 15 ؎ 2 kJ mol ؊1 and ⌬H 2 ‫؍‬ 8 ؎ 2 kJ mol ؊1

Surface‐Attached Polyhistidine‐Tag Proteins Characterized by FTIR Difference Spectroscopy

“…BeF 3 À mimics the g-phosphate and led to a conformational change of the GTPase from its "off" into its "on" state. [27] The observed difference spectrum is very similar to those obtained from Ras in solution and from lipidated membrane-bound Ras. [28] For example, the Thr35 marker band in the "on" state at 1684 cm À1 (in D 2 O) is well-resolved.…”