The N-terminal methyltransferase homologs NRMT1 and NRMT2 exhibit novel regulation of activity through heterotrimer formation.

protein;methylation;enzyme;epigenetic;regulation;structural

Jon David Faughn

University:University of Louisville

Department:Biochemistry and Molecular Biology

关键词: protein;    methylation;    enzyme;    epigenetic;    regulation;    structural;   

Others  :  https://ir.library.louisville.edu/cgi/viewcontent.cgi?article=4224&context=etd

美国|英语

来源: The Universite of Louisville's Institutional Repository

【 摘 要 】

Protein, DNA, and RNA methyltransferases have an ever-expanding list of novel substrates and catalytic activities. Even within families and between homologs, it is becoming clear the intricacies of methyltransferase specificity and regulation are far more diverse than originally thought. In addition to specific substrates and distinct methylation levels, methyltransferase activity can be altered through formation of complexes with close homologs. This work involves the N-terminal methyltransferase homologs NRMT1 and NRMT2. NRMT1 is a ubiquitously expressed distributive trimethylase. NRMT2 is a monomethylase expressed at low levels and in a tissue-specific manner. They are both nuclear methyltransferases with overlapping target consensus sequences but have distinct enzymatic activities and tissue expression patterns. Co-expression of NRMT1 with NRMT2 increases the trimethylation activity of NRMT1, and here I aim to understand how this occurs. I used analytical ultracentrifugation to show that while NRMT1 primarily exists as a dimer and NRMT2 as a monomer, when co-expressed they form a heterotrimer. I used co-immunoprecipitation and molecular modeling to demonstrate in vivo binding and map areas of interaction. While overexpression of NRMT2 increased the half-life of NRMT1, the reciprocal experiment did not produce the same results, indicating that NRMT2 may be increasing NRMT1 activity via increase the stability of the enzyme. Accordingly, the catalytic activity of NRMT2 is not needed to increase NRMT1 activity or increase its affinity for less preferred substrates. Additionally, monomethylation could not rescue phenotypes seen with loss of trimethylation. Taken together, these data support a model where NRMT2 expression activates NRMT1 activity, not through a catalytic change, but by increasing the stability and substrate affinity of NRMT1.