Substrate Specificity, Processivity, and Kinetic Mechanism
of Protein Arginine Methyltransferase 5
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Abstract
Protein arginine methyltransferases
(PRMTs) have emerged as attractive
therapeutic targets for heart disease and cancers. PRMT5 is a particularly
interesting target because it is overexpressed in blood, breast, colon,
and stomach cancers and promotes cell survival in the face of DNA
damaging agents. As the only known member of the PRMT enzyme family
to catalyze the formation of mono- and symmetrically dimethylated
arginine residues, PRMT5 is also mechanistically unique. As a part
of a program to characterize the mechanisms and regulation of the
PRMTs and develop chemical probes targeting these enzymes, we characterized
the substrate specificity, processivity, and kinetic mechanism of
bacterially expressed <i>Caenorhabditis elegans</i> PRMT5
(cPRMT5). In this report, we demonstrate that distal positively charged
residues contribute to substrate binding in a synergistic fashion.
Additionally, we show that cPRMT5 catalyzes symmetric dimethylation
in a distributive fashion. Finally, the results of initial velocity,
product, and dead-end inhibition studies indicate that cPRMT5 uses
a rapid equilibrium random mechanism with dead-end EAP and EBQ complexes.
In total, these studies will guide PRMT5 inhibitor development and
lay the foundation for studying how the activity of this medically
relevant enzyme is regulated