Biochemical Characterization of SET and MYND Domain Containing Protein 3

Abstract

The goal of this study is to reveal the biochemical function of SET and MYND domain containing protein 3 (SmyD3), a histone methyltransferase. Histone methylation is a vital epigenetic mechanism that controls many fundamental processes such as transcription activation and repression. SmyD3 methylates histone H3 at lysine 4 and was found greatly upregulated in several cancers including colon, liver, and breast cancers. However, the biochemical function of SMYD3 remains poorly understood. To gain insight into the function of SmyD3, we overexpressed mouse SmyD3 in E.coli cells. We found that SmyD3 elutes from a gel filtration column as a monomeric protein. Circular Dichroism (CD) spectroscopy showed that SmyD3 has a large amount of alpha‐helical structures. Thermal denaturation experiments showed that SmyD3 undergoes an unfolding transition with a melting temperature of ~ 45 °C. Our future studies will focus on that: first, understanding of how DNA binding to N‐terminal MYND domain impacts the second and tertiary structures of SmyD3 using Isothermal Titration Calorimetry and fluorescence spectroscopy; second, examination of the effects of substrate histone peptide binding on the protein structure and stability; finally, investigation of the function of the conserved C‐terminal domain, especially focusing on its contribution to protein integrity.