The PWWP domain of mammalian DNA methyltransferase Dnmt3b defines a new family of DNA-binding folds

Abstract

The PWWP domain is a weakly conserved sequence motif found in >60 eukaryotic proteins, including the mammalian DNA methyltransferases Dnmt3a and Dnmt3b. These proteins often contain other chromatin-association domains. A 135-residue PWWP domain from mouse Dnmt3b (amino acids 223-357) has been structurally characterized at 1.8 Å resolution. The N-terminal half of this domain resembles a barrel-like five-stranded structure, whereas the C-terminal half contains a five-helix bundle. The two halves are packed against each other to form a single structural module that exhibits a prominent positive electrostatic potential. The PWWP domain alone binds DNA in vitro, probably through its basic surface. We also show that recombinant Dnmt3b2 protein (a splice variant of Dnmt3b) and two N-terminal deletion mutants (Δ218 and Δ369) have approximately equal methyl transfer activity on unmethylated and hemimethylated CpG-containing oligonucleotides. The Δ218 protein, which includes the PWWP domain, binds DNA more strongly than Δ369, which lacks the PWWP domain.Genomic patterns of cytosine methylation at the C5 position play key roles in the organization and control of mammalian chromatin (reviewed in refs 1-3). Mammalian DNA methyltransferases (MTases) all contain a C-terminal catalytic region that is structurally and functionally homologous to bacterial MTases. The eukaryotic DNA MTases have been grouped into three families based on (i) distinctive properties of their N-terminal regions and (ii) intriguing differences in DNA substrate preferences. The first eukaryotic DNA MTase, Dnmt1, contains a large N-terminal regulatory region of ~1,000 amino acids and shows a preference for hemimethylated DNA in vitro [4][5][6] . Dnmt2, a relatively small protein, contains only the MTase domain, and its function is still unclear 7 . Recombinant Dnmt3 acts on unmethylated and hemimethylated DNA at equal rates in vitro 8,9 .The task of dissecting the functional roles of the N-terminal region of the different MTase families is just beginning. These regions vary widely in size and are probably responsible for the diverse biological functions of eukaryotic DNA MTases. These functions include the © 2002 Nature Publishing Group Correspondence should be addressed to X.C. xcheng@emory.edu. Competing interests statementThe authors declare that they have no competing financial interests. HHS Public Access Dnmt3 contains a PWWP domainThe sequences of Dnmt3 orthologs have been determined from human and mouse. They all contain an N-terminal variable region (~280 amino acids in Dnmt3a and ~220 amino acids in Dnmt3b), followed by three stretches of conserved regions: the PWWP motif, six repeats of a CXXC motif and a set of 10 motifs conserved among DNA MTase catalytic regions (Fig. 1a). The PWWP motif 28 was first identified in a gene family related to the hepatomaderived growth factor (HDGF) 29 and WHSC1 genes (Wolf-Hirschhorn Syndrome Candidate) 30 . The corresponding sequence in Dnmt3 is SWWP (Fig. 1b); only the last two positions of the motif...

Integrase, LEDGF/p75 and HIV replication

“…There are two dominant elements, a "PWWP domain" (residues 1-93) and an AT-Hook pair ( 178 PKRGRPAATEVKIPKPRGRPK 198 ). The PWWP domain is found in numerous other nuclear proteins [134,135]. Deletion of this domain or just its beta-barrel sub-domain from p75 disrupts microscopically detectable tethering to condensed chromosomes [93].…”

Diverse Domains of (Cytosine-5)-DNA Methyltransferases: Structural and Functional Characterization

“…However, it can also bind a nonspecific duplex of the same length with the same efficiency [117]. Deletion of the PWWP domain does not influence the Dnmt3b methylation efficiency in vitro [117]. On the contrary, the PWWP domain of Dnmt3a is almost unable to bind DNA [121].…”

“…The PWWP domain is named after a conserved ProTrpTrpPro motif, though the first Pro is substituted with Ser in Dnmt3a and Dnmt3b. The second part of the motif is always the same, TrpPro [117]. PWWP domain along with chromo domain, Tudor and MBT domains belongs to the Tudor domain "Royal family" [118,119].…”

“…The PWWP domain binds 30 bp duplexes with unmethylated, mono-, and dimethylated 5′-CG-3′/3′-GC-5′ sites. However, it can also bind a nonspecific duplex of the same length with the same efficiency [117]. Deletion of the PWWP domain does not influence the Dnmt3b methylation efficiency in vitro [117].…”

“…The PWWP domain of Dnmt3b has a positively charged surface with an approximate area of 45 × 32 Å 2 and can bind DNA nonspecifically [117,121]. The PWWP domain binds 30 bp duplexes with unmethylated, mono-, and dimethylated 5′-CG-3′/3′-GC-5′ sites.…”