X-ray structure of the complex of regulatory subunits of human DNA polymerase delta

The eukaryotic DNA polymerase delta (Pol delta) participates in genome replication, homologous recombination, DNA repair and damage tolerance. Regulation of the plethora of Pol delta functions depends on the interaction between the second (p50) and third (p66) non-catalytic subunits. We report the crystal structure of p50p66N complex featuring oligonucleotide binding and phosphodiesterase domains in p50 and winged helix-turn-helix N-terminal domain in p66. Disruption of the interaction between the yeast orthologs of p50 and p66 by strategic amino acid changes leads to cold-sensitivity, sensitivity to hydroxyurea and to reduced UV mutagenesis, mimicking the phenotypes of strains where the third subunit of Pol delta is absent. The second subunits of all B-family replicative DNA polymerases in archaea and eukaryotes, except Pol delta, share a three-domain structure similar to p50p66N, raising the possibility that a portion of the gene encoding p66 was derived from the second subunit gene relatively late in evolution.

[1]  T. Tahirov,et al.  Crystallization and preliminary crystallographic analysis of the complex of the second and third regulatory subunits of human Pol delta. , 2008, Acta Crystallographica. Section F : Structural Biology and Crystallization Communications.

[2]  Nick V. Grishin,et al.  Accurate statistical model of comparison between multiple sequence alignments , 2008, Nucleic acids research.

[3]  Michael J E Sternberg,et al.  Exploring the extremes of sequence/structure space with ensemble fold recognition in the program Phyre , 2008, Proteins.

[4]  T. Kunkel,et al.  The fidelity of DNA synthesis by eukaryotic replicative and translesion synthesis polymerases , 2008, Cell Research.

[5]  J. Haber,et al.  Break-induced replication and telomerase-independent telomere maintenance require Pol32 , 2007, Nature.

[6]  Tahir H. Tahirov,et al.  Structural basis for transcription elongation by bacterial RNA polymerase , 2007, Nature.

[7]  J. Prestegard,et al.  PF0610, a novel winged helix-turn-helix variant possessing a rubredoxin-like Zn ribbon motif from the hyperthermophilic archaeon, Pyrococcus furiosus. , 2007, Biochemistry.

[8]  D. Branzei,et al.  Mgs1 and Rad18/Rad5/Mms2 are required for survival of Saccharomyces cerevisiae mutants with novel temperature/cold sensitive alleles of the DNA polymerase delta subunit, Pol31. , 2006, DNA repair.

[9]  Marietta Y. W. T. Lee,et al.  Functional Roles of p12, the Fourth Subunit of Human DNA Polymerase δ* , 2006, Journal of Biological Chemistry.

[10]  Marietta Y. W. T. Lee,et al.  Direct Interaction of p21 with p50, the Small Subunit of Human DNA Polymerase Delta , 2006, Cell cycle.

[11]  I. Rogozin,et al.  Roles of DNA polymerases in replication, repair, and recombination in eukaryotes. , 2006, International review of cytology.

[12]  F. Asturias,et al.  Structure of Saccharomyces cerevisiae DNA polymerase epsilon by cryo–electron microscopy , 2006, Nature Structural &Molecular Biology.

[13]  C. Arrowsmith,et al.  Single-stranded DNA mimicry in the p53 transactivation domain interaction with replication protein A , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[14]  Emma Warbrick,et al.  An in vivo analysis of the localisation and interactions of human p66 DNA polymerase δ subunit , 2005, BMC Molecular Biology.

[15]  R. Woodgate,et al.  The Relative Roles in Vivo of Saccharomyces cerevisiae Pol η, Pol ζ, Rev1 Protein and Pol32 in the Bypass and Mutation Induction of an Abasic Site, T-T (6-4) Photoadduct and T-T cis-syn Cyclobutane Dimer , 2005, Genetics.

[16]  P. Burgers,et al.  DNA Polymerases that Propagate the Eukaryotic DNA Replication Fork , 2005, Critical reviews in biochemistry and molecular biology.

[17]  E. Warbrick,et al.  Mapping and mutation of the conserved DNA polymerase interaction motif (DPIM) located in the C-terminal domain of fission yeast DNA polymerase δ subunit Cdc27 , 2004, BMC Molecular Biology.

[18]  K Henrick,et al.  Electronic Reprint Biological Crystallography Secondary-structure Matching (ssm), a New Tool for Fast Protein Structure Alignment in Three Dimensions Biological Crystallography Secondary-structure Matching (ssm), a New Tool for Fast Protein Structure Alignment in Three Dimensions , 2022 .

[19]  Rosalind Kim,et al.  Structural and Functional Characterization of a Novel Phosphodiesterase from Methanococcus jannaschii* , 2004, Journal of Biological Chemistry.

[20]  L. F. Ciufo,et al.  The C-terminal zinc finger of the catalytic subunit of DNA polymerase delta is responsible for direct interaction with the B-subunit. , 2004, Nucleic acids research.

[21]  E. Johansson,et al.  The Pol32 Subunit of DNA Polymerase δ Contains Separable Domains for Processive Replication and Proliferating Cell Nuclear Antigen (PCNA) Binding* , 2004, Journal of Biological Chemistry.

[22]  W. Chazin,et al.  Independent and Coordinated Functions of Replication Protein A Tandem High Affinity Single-stranded DNA Binding Domains* , 2003, Journal of Biological Chemistry.

[23]  T. Kunkel,et al.  Roles of Saccharomyces cerevisiae DNA polymerases Poleta and Polzeta in response to irradiation by simulated sunlight. , 2003, Nucleic acids research.

[24]  J. Hurwitz,et al.  The Influence of the Cdc27 Subunit on the Properties of theSchizosaccharomyces pombe DNA Polymerase δ* , 2002, The Journal of Biological Chemistry.

[25]  C. Newlon,et al.  Yeast origins establish a strand bias for replicational mutagenesis. , 2002, Molecular cell.

[26]  C. Lawrence.  Cellular roles of DNA polymerase ζ and Rev1 protein , 2002 .

[27]  F. Galibert,et al.  Pol32, a subunit of Saccharomyces cerevisiae DNA polymerase delta, suppresses genomic deletions and is involved in the mutagenic bypass pathway. , 2002, Genetics.

[28]  G. Dianov,et al.  Reconstitution of human DNA polymerase delta using recombinant baculoviruses: the p12 subunit potentiates DNA polymerizing activity of the four-subunit enzyme. , 2002, The Journal of biological chemistry.

[29]  Takashi Kumasaka,et al.  Mechanism of c-Myb–C/EBPβ Cooperation from Separated Sites on a Promoter , 2002, Cell.

[30]  S. Ishii,et al.  Mechanism of c-Myb-C/EBP beta cooperation from separated sites on a promoter. , 2002, Cell.

[31]  T. Kunkel,et al.  In Vivo Consequences of Putative Active Site Mutations in Yeast DNA Polymerases α, ε, δ, and ζ , 2001 .

[32]  Thomas A. Steitz,et al.  Structure of the Replicating Complex of a Pol α Family DNA Polymerase , 2001, Cell.

[33]  H. Yoshikawa,et al.  The human homologue of fission Yeast cdc27, p66, is a component of active human DNA polymerase delta. , 2001, Journal of biochemistry.

[34]  F. Galibert,et al.  POL32, a subunit of the Saccharomyces cerevisiae DNA polymerase δ, defines a link between DNA replication and the mutagenic bypass repair pathway , 2000, Current Genetics.

[35]  C. Wittenberg,et al.  DNA Polymerase ε Catalytic Domains Are Dispensable for DNA Replication, DNA Repair, and Cell Viability , 1999 .

[36]  H. Pospiech,et al.  A novel family of DNA-polymerase-associated B subunits. , 1999, Trends in biochemical sciences.

[37]  F. Galibert,et al.  The Saccharomyces cerevisiae protein YJR043C (Pol32) interacts with the catalytic subunit of DNA polymerase α and is required for cell cycle progression in G2 / M , 1999, Molecular and General Genetics MGG.

[38]  R J Read,et al.  Crystallography & NMR system: A new software suite for macromolecular structure determination. , 1998, Acta crystallographica. Section D, Biological crystallography.

[39]  E V Koonin,et al.  Phosphoesterase domains associated with DNA polymerases of diverse origins. , 1998, Nucleic acids research.

[40]  P. Burgers,et al.  Characterization of the Two Small Subunits of Saccharomyces cerevisiae DNA Polymerase δ* , 1998, The Journal of Biological Chemistry.

[41]  Joël Janin,et al.  Specific versus non-specific contacts in protein crystals , 1997, Nature Structural Biology.

[42]  Alexey Bochkarev,et al.  Structure of the single-stranded-DNA-binding domain of replication protein A bound to DNA , 1997, Nature.

[43]  Gapped BLAST and PSI-BLAST: A new , 1997 .

[44]  S. Moreno,et al.  The fission yeast Cdc1 protein, a homologue of the small subunit of DNA polymerase delta, binds to Pol3 and Cdc27. , 1996, The EMBO journal.

[45]  Collaborative Computational,et al.  The CCP4 suite: programs for protein crystallography. , 1994, Acta crystallographica. Section D, Biological crystallography.

[46]  B. Stillman,et al.  Anatomy of a DNA replication fork revealed by reconstitution of SV40 DNA replication in vitro , 1994, Nature.

[47]  A. Murzin.  OB(oligonucleotide/oligosaccharide binding)‐fold: common structural and functional solution for non‐homologous sequences. , 1993, The EMBO journal.

[48]  K. Sharp,et al.  Protein folding and association: Insights from the interfacial and thermodynamic properties of hydrocarbons , 1991, Proteins.

[49]  C. Tan,et al.  DNA polymerase delta: A second eukaryotic DNA replicase , 1990, BioEssays : news and reviews in molecular, cellular and developmental biology.