Regulation of Protein Arginine Methyltransferase 8 (PRMT8) Activity by Its N-terminal Domain

Steven G. Clarke,Joyce Sayegh,Kristofor Webb,Donghang Cheng,Mark T. Bedford

doi:10.1074/jbc.m704650200

Abstract

Human protein arginine methyltransferase PRMT8 has been recently described as a type I enzyme in brain that is localized to the plasma membrane by N-terminal myristoylation. The amino acid sequence of human PRMT8 is almost 80% identical to human PRMT1, the major protein arginine methyltransferase activity in mammalian cells. However, the activity of a recombinant PRMT8 GST fusion protein toward methyl-accepting substrates is much lower than that of a GST fusion of PRMT1. We show here that both His-tagged and GST fusion species lacking the initial 60 amino acid residues of PRMT8 have enhanced enzymatic activity, suggesting that the N-terminal domain may regulate PRMT8 activity. This conclusion is supported by limited proteolysis experiments showing an increase in the activity of the digested full-length protein, consistent with the loss of the N-terminal domain. In contrast, the activity of the N-terminal truncated protein was slightly diminished by limited proteolysis. Significantly, we detect automethylation at two sites in the N-terminal domain, as well as binding sites for SH3 domain-containing proteins. We suggest that the N-terminal domain may function as an autoregulator that may be displaced by interaction with one or more physiological inducers.

Highlights

Methylation of arginine residues in proteins is catalyzed by four types of protein arginine methyltransferase enzymes (PRMTs).2 Type I, II, and III enzymes catalyze the transfer of
We find that the presence of the N-terminal region attenuates the activity of PRMT8 and suggest that the activity of this enzyme may be regulated by altering the conformation of the protein in this region
To find that the full-length N-terminal GST fusion of PRMT8 had much lower activity than the N-terminal GST fusion of PRMT1 when either GST-GAR (22) or myelin basic protein were used as methylaccepting substrates (Fig. 1)

Summary

Introduction

Methylation of arginine residues in proteins is catalyzed by four types of protein arginine methyltransferase enzymes (PRMTs).2 Type I, II, and III enzymes catalyze the transfer of. Digested GST-PRMT8 and GST-(⌬1– 60) PRMT8 (2 ␮g of protein) were incubated with 10 ␮g of GST-GAR and [3H]AdoMet (0.61 ␮M) at 37 °C for 1 h, followed by analysis of methylated products by SDS-PAGE and fluorography

Results

Conclusion