Inhibitory study of protein arginine methyltransferase 1 using a fluorescent approach

Biochem Biophys Res Commun. 2009 Feb 6;379(2):567-72. doi: 10.1016/j.bbrc.2008.12.119. Epub 2008 Dec 31.

Abstract

Protein arginine methyltransferases (PRMTs) play important roles in both normal physiology and human diseases. Deregulation of PRMT activity has been linked to several pathological states such as cancer and cardiovascular disorders. Herein, we report our work of designing and using new fluorescent reporters to perform single-step analysis of substrate binding and methylation by PRMT1. Both fluorescence intensity and anisotropy of the two reporters, R4-FL and H4-FL, were shown to effectively manifest enzyme-substrate interactions, highlighting their application in investigating PRMT inhibitors. In particular, the methylation process of R4-FL can be directly studied using fluorescence intensity readout. By combining the fluorescent measurement with radioactive analysis, we determined that AMI-1 inhibits PRMT1 activity through the mechanism of blocking peptide substrate binding.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Enzyme Inhibitors / pharmacology*
  • Fluorescence Polarization
  • Fluorescent Dyes / chemical synthesis
  • Fluorescent Dyes / chemistry*
  • Humans
  • Molecular Sequence Data
  • Naphthalenesulfonates / pharmacology*
  • Peptides / chemical synthesis
  • Peptides / chemistry*
  • Protein-Arginine N-Methyltransferases / antagonists & inhibitors*
  • Protein-Arginine N-Methyltransferases / chemistry
  • Repressor Proteins / antagonists & inhibitors*
  • Repressor Proteins / chemistry
  • Urea / analogs & derivatives*
  • Urea / pharmacology

Substances

  • 7,7'-carbonylbis(azanediyl) bis(4-hydroxynaphthalene-2-sulfonic acid
  • Enzyme Inhibitors
  • Fluorescent Dyes
  • Naphthalenesulfonates
  • Peptides
  • Repressor Proteins
  • Urea
  • PRMT1 protein, human
  • Protein-Arginine N-Methyltransferases