Targeting Ras palmitoylation for the treatment of cancer

Lay summary: Proteins can be thought of as molecular machines that carry out work within cells. Protein activity can be regulated through the reversible attachment of certain chemical groups to specific amino acids within proteins. Protein palmitoylation is one such modification and has been shown to control both the activity and location of proteins within mammalian cells. Protein palmitoylation regulates a wide range of processes within the body including how cells grow and divide. 20-30% of all cancers are caused by changes in the amino acid sequence of a protein called Ras. These changes cause Ras to be permanently active, which drives unregulated cell growth. To function correctly in the cell, Ras must be palmitoylated. Therefore, small molecule drugs that block Ras palmitoylation will likely be useful for treating cancer as they will block excess Ras activity preventing cells from growing and dividing. This Ph.D. is concerned with understanding the way that Ras is palmitoylated with a view to developing new drugs for the treatment of cancer.

Scientific summary: In humans, the genes HRAS, NRAS and KRAS encode three closely related proteins frequently mutated in cancer promoting unregulated cell growth. Ras proteins function as molecular switches transmitting signals from cell surface growth factor receptors to intracellular effector proteins cycling between inactive (GDP bound) and active (GTP) states. Oncogenic Ras activity requires the protein to be localised to the inner face of the plasma membrane suggesting strategies that cause Ras to be mistargeted within the cell may be effective in treating cancer. Ras membrane association is facilitated by two lipid post-translational modifications – farnesylation (all Ras isoforms) followed by palmitoylation (H-Ras, N-Ras and K-Ras4A only) – at specific cysteine residues located at the protein’s C-terminus. Ras is palmitoylated in the Golgi by an enzymatic complex consisting of zDHHC9 and GOLGA7. Molecules that bind to the surface of Ras and prevent it from interacting with the zDHHC9-GOLGA7 complex would be expected to selectively block Ras palmitoylation. In recent years, several groups have identified small molecules, peptides and Affimers that bind to shallow pockets on the surface of Ras blocking its activity. Yet, their effect on Ras palmitoylation has not been determined. In this Ph.D., the student will investigate the molecular basis of Ras palmitoylation using a range of biochemical, biophysical and pharmacological techniques. The novel, fundamental insights into Ras biology that will be gained are expected to open new drug discovery opportunities for the treatment of cancer.

Techniques: The student will gain experience of mammalian cell culture, molecular biology techniques (including PCR and site-directed mutagenesis), pharmacology, acyl-RAC (assay for determining protein palmitoylation levels), co-immunoprecipitation and pulldown experiments, SDS-PAGE and Western blotting, protein expression (bacteria, yeast) and purification as well as Hydrogen-Deuterium Exchange (HDX) mass spectrometry.