title: Activity and specificity of the human SUV39H2 protein lysine methyltransferase.
abstract: The SUV39H1 and SUV39H2 enzymes introduce H3K9me3, which is essential for the viability of mammalian cells. It was the aim of the present work to investigate the substrate specificity and product pattern of SUV39H2. Methylation of peptide SPOT arrays showed that SUV39H2 recognizes a long motif on H3 comprising T6-K14, with highly specific readout of R8, S10, T11 and G12 and partial specificity at T6, A7, G13 and K14. Modification of R8 and phosphorylation of S10 or T11 lead to a reduction or loss of SUV39H2 activity towards H3K9. The specificity of SUV39H2 differs from other H3K9 PKMTs, like Dim-5 or G9a, and these biochemical differences can be explained by the structures of the corresponding enzymes. Based on the specificity profile we identified additional non-histone candidate substrates in human proteins, but all of them were only weakly methylated by SUV39H2 at the peptide level. We conclude that SUV39H2 displays a high preference for the methylation of H3. Using the catalytic SET domain we show here that the enzyme prefers H3K9me0 as a substrate over H3K9me1 and H3K9me2 and it introduces the first two methyl groups into H3K9me0 in a processive reaction. SUV39H2 can transfer up to three methyl groups to lysine 9 of histone H3 but the last methylation reaction is much slower than the first two steps. We also demonstrate that the N324K mutant in the SET domain of SUV39H2 that has been shown to cause an inherited nasal skin disease in Labrador Retrievers renders SUV39H2 inactive. Differences in the circular dichroism spectra of wild type and mutant proteins indicated that the mutation causes slight structural changes.
authors.affiliation: Institute of Biochemistry, Stuttgart University, Pfaffenwaldring 55, 70569 Stuttgart, Germany.
authors.forename: Maren Kirstin
authors.initials: MK
authors.lastname: Schuhmacher
authors.affiliation: Institute of Biochemistry, Stuttgart University, Pfaffenwaldring 55, 70569 Stuttgart, Germany.
authors.forename: Srikanth
authors.initials: S
authors.lastname: Kudithipudi
authors.affiliation: Institute of Biochemistry, Stuttgart University, Pfaffenwaldring 55, 70569 Stuttgart, Germany.
authors.forename: Denis
authors.initials: D
authors.lastname: Kusevic
authors.affiliation: Institute of Biochemistry, Stuttgart University, Pfaffenwaldring 55, 70569 Stuttgart, Germany.
authors.forename: Sara
authors.initials: S
authors.lastname: Weirich
authors.affiliation: Institute of Biochemistry, Stuttgart University, Pfaffenwaldring 55, 70569 Stuttgart, Germany. Electronic address: albert.jeltsch@ibc.uni-stuttgart.de.
authors.forename: Albert
authors.initials: A
authors.lastname: Jeltsch
mesh_terms: D000595:Amino Acid Sequence; D002942:Circular Dichroism; D011495:Histone-Lysine N-Methyltransferase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D006657:Histones / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D008239:Lysine / Q000378:metabolism; D008745:Methylation; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics; D010766:Phosphorylation; D011487:Protein Conformation; D013379:Substrate Specificity
chemical_list: D006657:Histones; D050505:Mutant Proteins; D011495:Histone-Lysine N-Methyltransferase; C515701:SUV39H2 protein, human; D008239:Lysine
keywords: Enzyme specificity; H3K9 methylation; Heterochromatin; Histone post-translational modification; Lysine methylation