Raf kinases are downstream effectors of Ras proteins and are key players of the MAPK signaling pathway involved in a variety of cell signaling processes. Auto-inhibited cytosolic Raf is recruited to the plasma membrane by Ras for its normal function. Paradoxical activation is a serious problem in cancer patients where Raf-specific drug activates the MAPK signaling abnormally in a more aggressive manner. Majority of Raf-specific drugs target the ATP-binding pocket. Studies have demonstrated enhanced Raf dimerization by Raf-specific drugs and a predominant signaling via cRaf isoform of Raf (of three Raf kinases, a-, b- and cRaf) as primary causes that underlies the observed paradoxical activation. Disrupting Raf dimers will serve as a promising alternative strategy to abrogate abnormal MAPK signaling. My recent study based upon a sequence and structure-based bioinformatics analysis combined with microsecond-level all-atom molecular dynamics (MD) simulations revealed three distinct regions of the catalytic kinase domain of monomeric cRaf in direct contact with the membrane. cRaf dimer, on the other hand, although interacted with the membrane directly but transiently and peripherally as compared to the monomer. This was followed by performing probe-based MD simulation on the membrane-bound cRaf monomer and dimer (pMD-membrane). pMD is a methodology to identify small-molecule ligand-binding pockets in membrane-bound or water-soluble proteins. pMD was also performed on the monomer and dimers of cRaf in solution (pMD-solution). Binding free energy maps obtained from the pMD-membrane- and pMD-solution-derived conformational ensembles will be discussed with a focus on the cRaf dimer interface and may possibly yield potential ligand-binding pockets. To my knowledge, this is the first study that will utilize a membrane-dynamics guided conformational ensemble to probe for the potential ligand-binding pockets in Raf kinases.

Targeting membrane-bound dimer of cRaf kinase in search of anti-cancer drugs

2019 Spring National Meeting

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