In situ selectivity profiling and crystal structure of SML-8-73-1, an active site inhibitor of oncogenic K-Ras G12C

John C. Hunter, Deepak Gurbani, Scott B. Ficarro, Martin A. Carrasco, Sang Min Lim, Hwan Geun Choi, Ting Xie, Jarrod A. Marto, Zhe Chen, Nathanael S. Gray, Kenneth D. Westover

Research output: Contribution to journalArticle

107 Scopus citations

Abstract

Directly targeting oncogenic V-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (K-Ras) with small-molecule inhibitors has historically been considered prohibitively challenging. Recent reports of compounds that bind directly to the K-Ras G12C mutant suggest avenues to overcome key obstacles that stand in the way of developing such compounds. We aim to target the guanine nucleotide (GN)-binding pocket because the natural contents of this pocket dictate the signaling state of K-Ras. Here, we characterize the irreversible inhibitor SML-8-73-1 (SML), which targets the GN-binding pocket of K-Ras G12C. We report a high-resolution X-ray crystal structure of G12C K-Ras bound to SML, revealing that the compound binds in a manner similar to GDP, forming a covalent linkage with Cys-12. The resulting conformation renders K-Ras in the open, inactive conformation, which is not predicted to associate productively with or activate downstream effectors. Conservation analysis of the Ras family GN-binding pocket reveals variability in the side chains surrounding the active site and adjacent regions, especially in the switch I region. This variability may enable building specificity into new iterations of Ras and other GTPase inhibitors. High-resolution in situ chemical proteomic profiling of SML confirms that SML effectively discriminates between K-Ras G12C and other cellular GTP-binding proteins. A biochemical assay provides additional evidence that SML is able to compete with millimolar concentrations of GTP and GDP for the GN-binding site.

Original languageEnglish (US)
Pages (from-to)8895-8900
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume111
Issue number24
DOIs
Publication statusPublished - 2014

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Keywords

  • ActivX
  • CPM assay

ASJC Scopus subject areas

  • General

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