| Autor: |
Johanna M Jansen, Charles Wartchow, Wolfgang Jahnke, Susan Fong, Tiffany Tsang, Keith Pfister, Tatiana Zavorotinskaya, Dirksen Bussiere, Jan Marie Cheng, Kenneth Crawford, Yumin Dai, Jeffrey Dove, Eric Fang, Yun Feng, Jean-Michel Florent, John Fuller, Alvar D Gossert, Mohammad Hekmat-Nejad, Chrystèle Henry, Julia Klopp, William P Lenahan, Andreas Lingel, Sylvia Ma, Arndt Meyer, Yuji Mishina, Jamie Narberes, Gwynn Pardee, Savithri Ramurthy, Sebastien Rieffel, Darrin Stuart, Sharadha Subramanian, Laura Tandeske, Stephania Widger, Armin Widmer, Aurelie Winterhalter, Isabel Zaror, Stephen Hardy |
| Jazyk: |
angličtina |
| Rok vydání: |
2017 |
| Předmět: |
|
| Zdroj: |
PLoS ONE, Vol 12, Iss 4, p e0174706 (2017) |
| Druh dokumentu: |
article |
| ISSN: |
1932-6203 |
| DOI: |
10.1371/journal.pone.0174706 |
| Popis: |
RAS mutations lead to a constitutively active oncogenic protein that signals through multiple effector pathways. In this chemical biology study, we describe a novel coupled biochemical assay that measures activation of the effector BRAF by prenylated KRASG12V in a lipid-dependent manner. Using this assay, we discovered compounds that block biochemical and cellular functions of KRASG12V with low single-digit micromolar potency. We characterized the structural basis for inhibition using NMR methods and showed that the compounds stabilized the inactive conformation of KRASG12V. Determination of the biophysical affinity of binding using biolayer interferometry demonstrated that the potency of inhibition matches the affinity of binding only when KRAS is in its native state, namely post-translationally modified and in a lipid environment. The assays we describe here provide a first-time alignment across biochemical, biophysical, and cellular KRAS assays through incorporation of key physiological factors regulating RAS biology, namely a negatively charged lipid environment and prenylation, into the in vitro assays. These assays and the ligands we discovered are valuable tools for further study of KRAS inhibition and drug discovery. |
| Databáze: |
Directory of Open Access Journals |
| Externí odkaz: |
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