title: A kinetic study of human protein arginine N-methyltransferase 6 reveals a distributive mechanism.
abstract: Human protein arginine N-methyltransferase 6 (PRMT6) transfers methyl groups from the co-substrate S-adenosyl-L-methionine to arginine residues within proteins, forming S-adenosyl-L-homocysteine as well as omega-N(G)-monomethylarginine (MMA) and asymmetric dimethylarginine (aDMA) residues in the process. We have characterized the kinetic mechanism of recombinant His-tagged PRMT6 using a mass spectrometry method for monitoring the methylation of a series of peptides bearing a single arginine, MMA, or aDMA residue. We find that PRMT6 follows an ordered sequential mechanism in which S-adenosyl-L-methionine binds to the enzyme first and the methylated product is the first to dissociate. Furthermore, we find that the enzyme displays a preference for the monomethylated peptide substrate, exhibiting both lower K(m) and higher V(max) values than what are observed for the unmethylated peptide. This difference in substrate K(m) and V(max), as well as the lack of detectable aDMA-containing product from the unmethylated substrate, suggest a distributive rather than processive mechanism for multiple methylations of a single arginine residue. In addition, we speculate that the increased catalytic efficiency of PRMT6 for methylated substrates combined with lower K(m) values for native protein methyl acceptors may obscure this distributive mechanism to produce an apparently processive mechanism.
authors.affiliation: Division of Biomolecular & Pharmaceutical Chemistry, Faculty of Pharmaceutical Sciences, The University of British Columbia, Vancouver, British Columbia, Canada.
authors.forename: Ted M
authors.initials: TM
authors.lastname: Lakowski
authors.forename: Adam
authors.initials: A
authors.lastname: Frankel
mesh_terms: D001120:Arginine / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008745:Methylation; D009687:Nuclear Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding / Q000502:physiology; D011499:Protein Processing, Post-Translational / Q000502:physiology; D011484:Protein-Arginine N-Methyltransferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012435:S-Adenosylhomocysteine / Q000737:chemistry / Q000378:metabolism; D012436:S-Adenosylmethionine / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity / Q000502:physiology
chemical_list: D009687:Nuclear Proteins; D010455:Peptides; D011993:Recombinant Fusion Proteins; D012436:S-Adenosylmethionine; D001120:Arginine; D012435:S-Adenosylhomocysteine; C447833:PRMT6 protein, human; D011484:Protein-Arginine N-Methyltransferases