Activity and specificity of the human SUV39H2 protein lysine methyltransferase

Investigation of H2AX methylation by the SUV39H2 protein lysine methyltransferase

“…The sequence of the H3 tail perfectly fits to this profile. Our methylation studies with nonhistone candidate substrates showed the stringency of the peptide interaction of SUV39H2, because we identified three human proteins (Lysine‐specific demethylase 6A, WD repeat‐containing protein 43, and Zinc finger imprinted 2), which contain almost perfect target sites only differing from the SUV39H2 preferences at the + 4 and/or + 5 positions, but SUV39H2 could not methylate these peptides . Based on this, we compared the sequence surrounding lysine 134 in H2AX with the specificity profile of SUV39H2 and noticed that the H2AX residues at the − 1, + 1, + 3 and + 4 sites were not in agreement with the substrate recognition sequence motif of SUV39H2 (Fig.…”

“…H2AX (amino acids 118–143) was amplified from the cDNA clone obtained from the Dharmacon GE life sciences (Accession: , clone ID: 3139343) and cloned into the pGEX‐6p2 vector as GST fusion protein. The cloning of GST‐fused SUV39H2‐SET domain (amino acids 112–410) and His‐tagged full‐length SUV39H2 was described previously . GST‐fused murine Suv39h1 SET domain (82–412) cloned into pGEX‐2T vector was obtained as a gift from Dr Xiaodong Cheng.…”

“…However, SUV39H2 is highly expressed in various cancers such as lung cancers and acute lymphoblastic leukemia indicating that it is an important therapeutic target. The important role of SUV39H2 is further illustrated by the finding that somatic mutations in this (and other PKMTs) are observed in cancers and a hereditary disease in Labrador Retrievers has been mapped to a mutation in SUV39H2 which inactivates the enzyme .…”

Specificity of the SUV4–20H1 and SUV4–20H2 protein lysine methyltransferases and methylation of novel substrates

“…1). Such weak residual signals can be due to background binding of free or PKMT bound AdoMet to these peptide spots or they may reflect methylation of the cysteine residue present in the sequence as observed by us previously with the SET8 [34]. In short, we conclude that out of 7 peptide substrates tested SUV4-20H1 methylates one additional monomethylated substrate (CASZ1) and SUV4-20H2 methylates 3 novel peptide substrates (CASZ1, OIP5 and CENPU) indeed suggesting that SUV4-20H1 is more selective (as expected on the basis of the more specific peptide interaction) and that both enzymes differ in the spectrum of potential non-histone substrates.…”

“…For this purpose peptide libraries were synthesized using the SPOT synthesis method. This method has been successfully used to investigate the peptide interaction of different PKMTs previously (Dim5 [29], G9a [31], SET8 [18], SET7/9 [32], NSD1 [33], and SUV39H2 [34]). To study the substrate sequence motif of the SUV4-20H enzymes, peptide arrays were synthesized using the H4 (13-27) sequence with monomethylated K20 as template.…”