title: A System for Enzymatic Lysine Methylation in a Desired Sequence Context.
abstract: A number of lysine-specific methyltransferases (KMTs) are responsible for the post-translational modification of cellular proteins on lysine residues. Most KMTs typically recognize specific motifs in unstructured, short peptide sequences. However, we have recently discovered a novel KMT that appeared to have a more relaxed sequence specificity, namely, valosin-containing protein (VCP)-KMT, which trimethylates Lys-315 in the molecular chaperone VCP. On the basis of this, here, we explored the possibility of using the VCP-KMT/VCP system to obtain specific lysine methylation of desired sequences grafted onto a VCP-derived scaffold. We generated VCP-derived proteins in which three amino acid residues on each side of Lys-315 had been replaced by various sequences representing lysine methylation sites in histone H3. We found that all of these chimeric proteins were subject to efficient VCP-KMT-mediated methylation in vitro, and methylation was also observed in mammalian cells. Thus, we here describe a versatile system for introducing lysine methylation into a desired peptide sequence, and the approach should be readily expandable for generating combinatorial libraries of methylated sequences.
authors.affiliation: Department of Biosciences, University of Oslo , P.O. Box 1066, Blindern, 0316 Oslo, Norway.
authors.forename: Vinay Kumar
authors.initials: VK
authors.lastname: Aileni
authors.affiliation: Department of Biosciences, University of Oslo , P.O. Box 1066, Blindern, 0316 Oslo, Norway.
authors.forename: Erna
authors.initials: E
authors.lastname: Davydova
authors.affiliation: Department of Biosciences, University of Oslo , P.O. Box 1066, Blindern, 0316 Oslo, Norway.
authors.forename: Anders
authors.initials: A
authors.lastname: Moen
authors.affiliation: Department of Biosciences, University of Oslo , P.O. Box 1066, Blindern, 0316 Oslo, Norway.
authors.forename: Pål Ø
authors.initials: PØ
authors.lastname: Falnes