Dataset Information

Insights into the Cross Talk between Effector and Allosteric Lobes of KRAS from Methyl Conformational Dynamics.

ABSTRACT: KRAS is the most frequently mutated RAS protein in cancer patients, and it is estimated that about 20% of the cancer patients in the United States carried mutant RAS proteins. To accelerate therapeutic development, structures and dynamics of RAS proteins had been extensively studied by various biophysical techniques for decades. Although ³¹P NMR studies revealed population equilibrium of the two major states in the active GMPPNP-bound form, more complex conformational dynamics in RAS proteins and oncogenic mutants subtly modulate the interactions with their downstream effectors. We established a set of customized NMR relaxation dispersion techniques to efficiently and systematically examine the ms-μs conformational dynamics of RAS proteins. This method allowed us to observe varying synchronized motions that connect the effector and allosteric lobes in KRAS. We demonstrated the role of conformational dynamics of KRAS in controlling its interaction with the Ras-binding domain of the downstream effector RAF1, the first kinase in the MAPK pathway. This allows one to explain, as well as to predict, the altered binding affinities of various KRAS mutants, which was neither previously reported nor apparent from the structural perspective.

SUBMITTER: Chao FA

PROVIDER: S-EPMC10430694 | biostudies-literature | 2022 Mar

REPOSITORIES: biostudies-literature

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Insights into the Cross Talk between Effector and Allosteric Lobes of KRAS from Methyl Conformational Dynamics.

Chao Fa-An FA Dharmaiah Srisathiyanarayanan S Taylor Troy T Messing Simon S Gillette William W Esposito Dominic D Nissley Dwight V DV McCormick Frank F Byrd R Andrew RA Simanshu Dhirendra K DK Cornilescu Gabriel G

Journal of the American Chemical Society 20220225 9

KRAS is the most frequently mutated RAS protein in cancer patients, and it is estimated that about 20% of the cancer patients in the United States carried mutant RAS proteins. To accelerate therapeutic development, structures and dynamics of RAS proteins had been extensively studied by various biophysical techniques for decades. Although <sup>31</sup>P NMR studies revealed population equilibrium of the two major states in the active GMPPNP-bound form, more complex conformational dynamics in RAS ...[more]

PMID: 35213144

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Project description:Calmodulin is a calcium-binding protein ubiquitous in eukaryotic cells, involved in numerous calcium-regulated biological phenomena, such as synaptic plasticity, muscle contraction, cell cycle, and circadian rhythms. It exibits a characteristic dumbell shape, with two globular domains (N- and C-terminal lobe) joined by a linker region. Each lobe can take alternative conformations, affected by the binding of calcium and target proteins. Calmodulin displays considerable functional flexibility due to its capability to bind different targets, often in a tissue-specific fashion. In various specific physiological environments (e.g. skeletal muscle, neuron dendritic spines) several targets compete for the same calmodulin pool, regulating its availability and affinity for calcium. In this work, we sought to understand the general principles underlying calmodulin modulation by different target proteins, and to account for simultaneous effects of multiple competing targets, thus enabling a more realistic simulation of calmodulin-dependent pathways. We built a mechanistic allosteric model of calmodulin, based on an hemiconcerted framework: each calmodulin lobe can exist in two conformations in thermodynamic equilibrium, with different affinities for calcium and different affinities for each target. Each lobe was allowed to switch conformation on its own. The model was parameterised and validated against experimental data from the literature. In spite of its simplicity, a two-state allosteric model was able to satisfactorily represent several sets of experiments, in particular the binding of calcium on intact and truncated calmodulin and the effect of different skMLCK peptides on calmodulin's saturation curve. The model can also be readily extended to include multiple targets. We show that some targets stabilise the low calcium affinity T state while others stabilise the high affinity R state. Most of the effects produced by calmodulin targets can be explained as modulation of a pre-existing dynamic equilibrium between different conformations of calmodulin's lobes, in agreement with linkage theory and MWC-type models.

Dataset Information

Insights into the Cross Talk between Effector and Allosteric Lobes of KRAS from Methyl Conformational Dynamics.

Publications

Insights into the Cross Talk between Effector and Allosteric Lobes of KRAS from Methyl Conformational Dynamics.

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