The Importance of Mg(2+)‐Free State in Nucleotide Exchange of Oncogenic K‐Ras Mutants

For efficient targeting of oncogenic K‐Ras interaction sites, a mechanistic picture of the Ras‐cycle is necessary. Herein, we used NMR relaxation techniques and molecular dynamics simulations to decipher the role of slow dynamics in wild‐type and three oncogenic P‐loop mutants of K‐Ras. Our measurements reveal a dominant two‐state conformational exchange on the ms timescale in both GDP‐ and GTP‐bound K‐Ras. The identified low‐populated higher energy state in GDP‐loaded K‐Ras has a conformation reminiscent of a nucleotide‐bound/Mg(2+)‐free state characterized by shortened β2/β3‐strands and a partially released switch‐I region preparing K‐Ras for the interaction with the incoming nucleotide exchange factor and subsequent reactivation. By providing insight into mutation‐specific differences in K‐Ras structural dynamics, our systematic analysis improves our understanding of prolonged K‐Ras signaling and may aid the development of allosteric inhibitors targeting nucleotide exchange in K‐Ras.