Functional Characterization of SET domain protein eN-methyltransferases

Elizabeth Megan Flynn
University of Kentucky

Rubisco LSMT is a SET (Suppressor of variegation 3-9, Enhancer of zeste, Trithorax) domain protein methyltransferase responsible for the post-translational formation of trimethyllysyl residues at position 14 in the N-terminal region of the large subunit of Rubisco. In contrast to many SET domain histone specific methyltransferases, Rubisco LSMT also contains a structurally unique C-terminal domain comprised of 176 amino acid residues. Based on models mimicking the binding between Rubisco LSMT and Rubisco, the C-terminal domain may contribute to the specificity and interaction between these two proteins. Also, expression of N-terminally truncated forms of Rubisco LSMT and subsequent binding analysis using ELISA suggest that the C-terminal domain can interact with Rubisco in the absence of the N-terminal catalytic SET domain. To examine the role that the C-terminal domain plays in catalysis as well as substrate binding we are expressing truncated constructs of Rubisco LSMT. Furthermore, we are investigating homologous ORFs from humans and mice for analysis of protein methyltransferase activity and the role, if any, that the C-terminal domain plays in these proteins. The function of the C-terminal in the interaction of Rubisco LSMT with Rubisco may have important implications for other protein methyltransferases given that this sequence is found with 80% consensus in a number of ORFs from a wide range of species including Neurospora crassa, Drosophila melongaster, Bos taurus, and Sus scrofa.