title: Structural insights into recognition of MDC1 by TopBP1 in DNA replication checkpoint control.
abstract: Activation of the DNA replication checkpoint by the ATR kinase requires protein interactions mediated by the ATR-activating protein, TopBP1. Accumulation of TopBP1 at stalled replication forks requires the interaction of TopBP1 BRCT5 with the phosphorylated SDT repeats of the adaptor protein MDC1. Here, we present the X-ray crystal structures of the tandem BRCT4/5 domains of TopBP1 free and in complex with a MDC1 consensus pSDpT phosphopeptide. TopBP1 BRCT4/5 adopts a variant BRCT-BRCT packing interface and recognizes its target peptide in a manner distinct from that observed in previous tandem BRCT- peptide structures. The phosphate-binding pocket and positively charged residues in a variant loop in BRCT5 present an extended binding surface for the negatively charged MDC1 phosphopeptide. Mutations in this surface reduce binding affinity and recruitment of TopBP1 to γH2AX foci in cells. These studies reveal a different mode of phosphopeptide binding by BRCT domains in the DNA damage response.
authors.affiliation: Department of Biochemistry, University of Alberta, Edmonton, Alberta T6G 2H7, Canada.
authors.forename: Charles Chung Yun
authors.initials: CC
authors.lastname: Leung
authors.forename: Luxin
authors.initials: L
authors.lastname: Sun
authors.forename: Zihua
authors.initials: Z
authors.lastname: Gong
authors.forename: Michael
authors.initials: M
authors.lastname: Burkat
authors.forename: Ross
authors.initials: R
authors.lastname: Edwards
authors.forename: Mark
authors.initials: M
authors.lastname: Assmus
authors.forename: Junjie
authors.initials: J
authors.lastname: Chen
authors.forename: J N Mark
authors.initials: JN
authors.lastname: Glover
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001665:Binding Sites; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics; D018797:Cell Cycle Proteins; D018360:Crystallography, X-Ray; D042522:DNA Replication Timing; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics; D006801:Humans; D006860:Hydrogen Bonding; D057927:Hydrophobic and Hydrophilic Interactions; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D009687:Nuclear Proteins / Q000737:chemistry* / Q000235:genetics; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D020449:Repetitive Sequences, Amino Acid; D015534:Trans-Activators / Q000737:chemistry*
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C413163:MDC1 protein, human; D009687:Nuclear Proteins; C110513:TOPBP1 protein, human; D015534:Trans-Activators
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