A Comparative CEST NMR Study of Slow Conformational Dynamics of Small GTPases Complexed with GTP and GTP Analogues

Abstract

The Ras superfamily of small GTPases are important intracellular signaling molecules, the functions of which are determined by the binding of guanosine nucleotides (GTP = guanosine triphosphate and GDP = guanosine diphosphate). [1] The GTP-bound (“active”) states of these enzymes are capable of interacting with specific downstream effector proteins, thus eliciting a wide range of cellular responses. [2, 3] Mutations that reduce the rate of GTP hydrolysis and thus increase the lifetime of the active GTP-bound state are frequently oncogenic and contribute to the development and metastasis of human cancers. [4] Elegant 31 P NMR studies of GTP-bound Ras showed that the enzyme interconverts between two states, a minor conformer termed state 1 and a major species designated state 2. [5–9] Similar conformational dynamics have been observed in other Ras family GTPases as well. [10–12] State 2 is generally regarded as the conformation competent for binding effector proteins, whereas state 1 exhibits significantly reduced affinity for these molecules. [5–7, 13, 14] Stabilization of the low-affinity state 1 was