Advertisement

Extremophiles

, Volume 23, Issue 1, pp 161–172 | Cite as

Elucidating functions of DP1 and DP2 subunits from the Thermococcus kodakarensis family D DNA polymerase

  • Natsuki Takashima
  • Sonoko IshinoEmail author
  • Keisuke Oki
  • Mika Takafuji
  • Takeshi Yamagami
  • Ryotaro Matsuo
  • Kouta Mayanagi
  • Yoshizumi IshinoEmail author
Original Paper

Abstract

DNA polymerase D (PolD), originally discovered in Pyrococcus furiosus, has no sequence homology with any other DNA polymerase family. Genes encoding PolD are found in most of archaea, except for those archaea in the Crenarchaeota phylum. PolD is composed of two proteins: DP1 and DP2. To date, the 3D structure of the PolD heteromeric complex is yet to be determined. In this study, we established a method that prepared highly purified PolD from Thermococcus kodakarensis, and purified DP1 and DP2 proteins formed a stable complex in solution. An intrinsically disordered region was identified in the N-terminal region of DP1, but the static light scattering analysis provided a reasonable molecular weight of DP1. In addition, PolD forms as a complex of DP1 and DP2 in a 1:1 ratio. Electron microscope single particle analysis supported this composition of PolD. Both proteins play an important role in DNA synthesis activity and in 3′–5′ degradation activity. DP1 has extremely low affinity for DNA, while DP2 is mainly responsible for DNA binding. Our work will provide insight and the means to further understand PolD structure and the molecular mechanism of this archaea-specific DNA polymerase.

Keywords

Thermococcus kodakarensis DNA polymerase DNA replication Archaea Euryarchaeota 

Abbreviations

SLS

Static light scattering

CD

Circular dichroism

EM

Electron microscopy

RALS

Right angle light scattering

Notes

Acknowledgements

The authors thank Ryo Ugawa (Research Promotion Unit, Medical Institute of Bioregulation, Kyushu University) for technical assistance.

Funding

This work was supported by grants from the Ministry of Education, Culture, Sports, Science and Technology of Japan (Grant numbers JP21113005, JP23310152, and JP26242075 to Y. I., JP16H01410 to K.M., and JP18K05442 to S.I.). K.M. was also supported by JST PRESTO (JPMJPR12L9)

Compliance with ethical standards

Conflict of interest

None declared.

Supplementary material

792_2018_1070_MOESM1_ESM.pdf (3.3 mb)
Supplementary material 1 (PDF 3416 kb)

References

  1. Abellon-Ruiz J, Waldron KJ, Connolly BA (2016) Archaeoglobus fulgidus DNA polymerase D: a zinc-binding protein inhibited by hypoxanthine and uracil. J Mol Biol 428:2805–2813.  https://doi.org/10.1016/j.jmb.2016.06.008 CrossRefGoogle Scholar
  2. Berquist BR, DasSarma P, DasSarma S (2007) Essential and non-essential DNA replication genes in the model halophilic Archaeon, Halobacterium sp. NRC-1. BMC Genet 8:31.  https://doi.org/10.1186/1471-2156-8-31 CrossRefGoogle Scholar
  3. Cann IK, Ishino Y (1999) Archaeal DNA replication: identifying the pieces to solve a puzzle. Genetics 152:1249–1267Google Scholar
  4. Cann IK, Komori K, Toh H, Kanai S, Ishino Y (1998) A heterodimeric DNA polymerase: evidence that members of Euryarchaeota possess a distinct DNA polymerase. Proc Natl Acad Sci USA 95:14250–14255CrossRefGoogle Scholar
  5. Castrec B, Laurent S, Henneke G, Flament D, Raffin JP (2010) The glycine-rich motif of Pyrococcus abyssi DNA polymerase D is critical for protein stability. J Mol Biol 396:840–848.  https://doi.org/10.1016/j.jmb.2010.01.006 CrossRefGoogle Scholar
  6. Cubonova L, Richardson T, Burkhart BW, Kelman Z, Connolly BA, Reeve JN, Santangelo TJ (2013) Archaeal DNA polymerase D but not DNA polymerase B is required for genome replication in Thermococcus kodakarensis. J Bacteriol 195:2322–2328.  https://doi.org/10.1128/JB.02037-12 CrossRefGoogle Scholar
  7. Gasteiger E, Gattiker A, Hoogland C, Ivanyi I, Appel RD, Bairoch A (2003) ExPASy: the proteomics server for in-depth protein knowledge and analysis. Nucleic Acids Res 31:3784–3788.  https://doi.org/10.1093/nar/gkg563 CrossRefGoogle Scholar
  8. Greenough L, Menin JF, Desai NS, Kelman Z, Gardner AF (2014) Characterization of family D DNA polymerase from Thermococcus sp. 9 degrees N. Extremophiles 18:653–664.  https://doi.org/10.1007/s00792-014-0646-9 CrossRefGoogle Scholar
  9. Greenough L, Kelman Z, Gardner AF (2015) The roles of family B and D DNA polymerases in Thermococcus species 9 degrees N Okazaki fragment maturation. J Biol Chem 290:12514–12522.  https://doi.org/10.1074/jbc.M115.638130 CrossRefGoogle Scholar
  10. Gueguen Y et al (2001) Characterization of two DNA polymerases from the hyperthermophilic euryarchaeon Pyrococcus abyssi. Eur J Biochem 268:5961–5969CrossRefGoogle Scholar
  11. Henneke G, Flament D, Hubscher U, Querellou J, Raffin JP (2005) The hyperthermophilic euryarchaeota Pyrococcus abyssi likely requires the two DNA polymerases D and B for DNA replication. J Mol Biol 350:53–64.  https://doi.org/10.1016/j.jmb.2005.04.042 CrossRefGoogle Scholar
  12. Imamura M, Uemori T, Kato I, Ishino Y (1995) A non-alpha-like DNA polymerase from the hyperthermophilic archaeon Pyrococcus furiosus. Biol Pharm Bull 18:1647–1652.  https://doi.org/10.1248/bpb.18.1647 CrossRefGoogle Scholar
  13. Ishino S, Ishino Y (2006) Comprehensive search for DNA polymerase in the hyperthermophilic archaeon, Pyrococcus furiosus. Nucleosides Nucleotides Nucleic Acids 25:681–691CrossRefGoogle Scholar
  14. Ishino Y, Ishino S (2012) Rapid progress of DNA replication studies in archaea, the third domain of life. Sci China Life Sci 55:386–403.  https://doi.org/10.1007/s11427-012-4324-9 CrossRefGoogle Scholar
  15. Ishino Y, Komori K, Cann IK, Koga Y (1998) A novel DNA polymerase family found in Archaea. J Bacteriol 180:2232–2236Google Scholar
  16. Jokela M, Eskelinen A, Pospiech H, Rouvinen J, Syvaoja JE (2004) Characterization of the 3′ exonuclease subunit DP1 of Methanococcus jannaschii replicative DNA polymerase D. Nucleic Acids Res 32:2430–2440.  https://doi.org/10.1093/nar/gkh558 CrossRefGoogle Scholar
  17. Kimanius D, Forsberg BO, Scheres SH, Lindahl E (2016) Accelerated cryo-EM structure determination with parallelisation using GPUs in RELION-2. Elife.  https://doi.org/10.7554/elife.18722 Google Scholar
  18. Komori K, Ichiyanagi K, Morikawa K, Ishino Y (1999) PI-PfuI and PI-PfuII, intein-coded homing endonucleases from Pyrococcus furiosus. II. Characterization Of the binding and cleavage abilities by site-directed mutagenesis. Nucleic Acids Res 27:4175–4182.  https://doi.org/10.1093/nar/27.21.417 CrossRefGoogle Scholar
  19. Kuba Y et al (2012) Comparative analyses of the two proliferating cell nuclear antigens from the hyperthermophilic archaeon, Thermococcus kodakarensis. Genes Cells 17:923–937.  https://doi.org/10.1111/gtc.12007 CrossRefGoogle Scholar
  20. Makarova KS, Krupovic M, Koonin EV (2014) Evolution of replicative DNA polymerases in archaea and their contributions to the eukaryotic replication machinery. Front Microbiol 5:354.  https://doi.org/10.3389/fmicb.2014.00354 CrossRefGoogle Scholar
  21. Masai H, Matsumoto S, You Z, Yoshizawa-Sugata N, Oda M (2010) Eukaryotic chromosome DNA replication: where, when, and how? Annu Rev Biochem 79:89–130.  https://doi.org/10.1146/annurev.biochem.052308.103205 CrossRefGoogle Scholar
  22. Matsui I, Urushibata Y, Shen Y, Matsui E, Yokoyama H (2011) Novel structure of an N-terminal domain that is crucial for the dimeric assembly and DNA-binding of an archaeal DNA polymerase D large subunit from Pyrococcus horikoshii. FEBS Lett 585:452–458.  https://doi.org/10.1016/j.febslet.2010.12.040 CrossRefGoogle Scholar
  23. Matsui I, Matsui E, Yamasaki K, Yokoyama H (2013) Domain structures and inter-domain interactions defining the holoenzyme architecture of archaeal D-family DNA polymerase. Life (Basel) 3:375–385.  https://doi.org/10.3390/life3030375 Google Scholar
  24. Mott ML, Berger JM (2007) DNA replication initiation: mechanisms and regulation in bacteria. Nat Rev Microbiol 5:343–354.  https://doi.org/10.1038/nrmicro1640 CrossRefGoogle Scholar
  25. Nagata M et al (2017) The Cdc45/RecJ-like protein forms a complex with GINS and MCM, and is important for DNA replication in Thermococcus kodakarensis. Nucleic Acids Res 45:10693–10705.  https://doi.org/10.1093/nar/gkx740 CrossRefGoogle Scholar
  26. Ollis DL, Brick P, Hamlin R, Xuong NG, Steitz TA (1985) Structure of large fragment of Escherichia coli DNA polymerase I complexed with dTMP. Nature 313:762–766CrossRefGoogle Scholar
  27. Pettersen EF, Goddard TD, Huang CC, Couch GS, Greenblatt DM, Meng EC, Ferrin TE (2004) UCSF Chimera—a visualization system for exploratory research and analysis. J Comput Chem 25:1605–1612.  https://doi.org/10.1002/jcc.20084 CrossRefGoogle Scholar
  28. Sarmiento F, Mrazek J, Whitman WB (2013) Genome-scale analysis of gene function in the hydrogenotrophic methanogenic archaeon Methanococcus maripaludis. Proc Natl Acad Sci USA 110:4726–4731.  https://doi.org/10.1073/pnas.1220225110 CrossRefGoogle Scholar
  29. Sauguet L, Raia P, Henneke G, Delarue M (2016) Shared active site architecture between archaeal PolD and multi-subunit RNA polymerases revealed by X-ray crystallography. Nat Commun 7:12227.  https://doi.org/10.1038/ncomms12227 CrossRefGoogle Scholar
  30. Schermerhorn KM, Gardner AF (2015) Pre-steady-state kinetic analysis of a family D DNA polymerase from Thermococcus sp. 9 degrees N reveals mechanisms for archaeal genomic replication and maintenance. J Biol Chem 290:21800–21810.  https://doi.org/10.1074/jbc.M115.662841 CrossRefGoogle Scholar
  31. Shen Y, Musti K, Hiramoto M, Kikuchi H, Kawarabayashi Y, Matsui I (2001) Invariant Asp-1122 and Asp-1124 are essential residues for polymerization catalysis of family D DNA polymerase from Pyrococcus horikoshii. J Biol Chem 276:27376–27383.  https://doi.org/10.1074/jbc.M011762200 CrossRefGoogle Scholar
  32. Shen Y, Tang XF, Matsui I (2003) Subunit interaction and regulation of activity through terminal domains of the family D DNA polymerase from Pyrococcus horikoshii. J Biol Chem 278:21247–21257.  https://doi.org/10.1074/jbc.M212286200 CrossRefGoogle Scholar
  33. Shen Y, Tang XF, Yokoyama H, Matsui E, Matsui I (2004) A 21-amino acid peptide from the cysteine cluster II of the family D DNA polymerase from Pyrococcus horikoshii stimulates its nuclease activity which is Mre11-like and prefers manganese ion as the cofactor. Nucleic Acids Res 32:158–168.  https://doi.org/10.1093/nar/gkh153 CrossRefGoogle Scholar
  34. Tang XF, Shen Y, Matsui E, Matsui I (2004) Domain topology of the DNA polymerase D complex from a hyperthermophilic archaeon Pyrococcus horikoshii. Biochemistry 43:11818–11827.  https://doi.org/10.1021/bi0362931 CrossRefGoogle Scholar
  35. Uemori T, Ishino Y, Toh H, Asada K, Kato I (1993) Organization and nucleotide sequence of the DNA polymerase gene from the archaeon Pyrococcus furiosus. Nucleic Acids Res 21:259–265CrossRefGoogle Scholar
  36. Uemori T, Sato Y, Kato I, Doi H (1997) A novel DNA polymerase in the hyperthermophilic archaeon, Pyrococcus furiosus: gene cloning, expression, and characterization. Genes Cells 2:499–512CrossRefGoogle Scholar
  37. Uversky VN (2012) Size-exclusion chromatography in structural analysis of intrinsically disordered proteins. Methods Mol Biol 896:179–194.  https://doi.org/10.1007/978-1-4614-3704-8_11 Google Scholar
  38. Yamasaki K, Urushibata Y, Yamasaki T, Arisaka F, Matsui I (2010) Solution structure of the N-terminal domain of the archaeal D-family DNA polymerase small subunit reveals evolutionary relationship to eukaryotic B-family polymerases. FEBS Lett 584:3370–3375.  https://doi.org/10.1016/j.febslet.2010.06.026 CrossRefGoogle Scholar
  39. Zhang K (2016) Gctf: real-time CTF determination and correction. J Struct Biol 193:1–12.  https://doi.org/10.1016/j.jsb.2015.11.003 CrossRefGoogle Scholar

Copyright information

© Springer Japan KK, part of Springer Nature 2018

Authors and Affiliations

  • Natsuki Takashima
    • 1
  • Sonoko Ishino
    • 1
    Email author
  • Keisuke Oki
    • 1
  • Mika Takafuji
    • 1
  • Takeshi Yamagami
    • 1
  • Ryotaro Matsuo
    • 3
  • Kouta Mayanagi
    • 2
  • Yoshizumi Ishino
    • 1
    Email author
  1. 1.Graduate School of Bioresource and Bioenvironmental SciencesKyushu UniversityFukuokaJapan
  2. 2.Medical Institute of BioregulationKyushu UniversityFukuokaJapan
  3. 3.Division of Malvern PanalyticalSpectris Co., LtdTokyoJapan

Personalised recommendations