UCLA

Methylation is an essential post translational modification that can be found on a variety of proteins in higher and lower species. The effects of these modifications are diverse and include targeting genes for activation or silencing and protein stability. The most extensively characterized methylation reaction is that of histone tails. In the past decade researchers became interested in identifying non-histone protein substrates and the enzymes responsible for methylating them (methyltransferases). In the yeast Saccharomyces cerevisiae, a substantial number of methyltransferases have been identified for modifying mitochondrial proteins, translational release factors, ribosomal proteins and translational elongation factors.

When I started my doctoral journey the newest methyltransferase of the yeast translational apparatus was found to modify elongation factor 1 alpha (EF1A), the protein responsible for bringing the aminoacyl-tRNA to the ribosome. This enzyme, Efm7, was the fifth elongation factor methyltransferase (Efm) responsible for methylating EF1A! EF1A methylation by five methyltransferases is unique as other elongation factors, EF2 and EF3, only have two methyltransferases that modify them both. Additionally, each of these five EF1A methyltransferases appears to target specific residues on EF1A. This has not been seen in any other protein of the translational apparatus that is methylated. Thus I was captivated by this uniqueness of EF1A methylation.

Although it is known that EF1A and other translational proteins are methylated and the enzymes responsible for its methylation, we still don’t know why it is methylated. GTP-bound EF1A functions as the courier of aminoacylated tRNA to the ribosome. When a correct codon-anticodon match is made, a conformational change in the ribosome results in the hydrolysis of GTP to GDP and the release of EF1A. Additionally, GDP bound-EF1A functions outside of the ribosome including being able to bind and bundle filamentous actin. This dissertation focused on identifying whether or not there was a direct link between methylation and functionality of EF1A. In a separate study, I examined the functional role of ribosomal protein methyltransferases.

My studies lead to the development and characterization of the first quintuplet knockout yeast strain for the enzymes responsible for methylating EF1A. These strains have been confirmed to be methylation deficient at the respective EF1A lysine sites, using mass spectrometry. I found that EF1A methylation appears to be necessary for helping yeast to survive and adapt to differences in its cellular environment but may not be necessary for the integrity of the actin cytoskeleton as well as EF1A induced actin binding.