Biomolecular NMR Assignments

, Volume 12, Issue 1, pp 107–111 | Cite as

Backbone 1H, 13C and 15N resonance assignments of the OB domain of the single stranded DNA-binding protein hSSB2 (NABP1/OBFC2A) and chemical shift mapping of the DNA-binding interface

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Abstract

Single stranded DNA-binding proteins (SSBs) are essential for the maintenance of genome integrity and are required in in all known cellular organisms. Over the last 10 years, the role of two new human SSBs, hSSB1 (NABP2/OBFC2B) and hSSB2 (NABP1/OBFC2A), has been described and characterised in various important DNA repair processes. Both these proteins are made up of a conserved oligonucleotide-binding (OB) fold that is responsible for ssDNA recognition as well a unique flexible carboxy-terminal extension involved in protein–protein interactions. Due to their similar domain organisation, hSSB1 and hSSB2 have been found to display some overlapping functions. However, several studies have also revealed cell- and tissue-specific roles for these two proteins, most likely due to small but significant differences in the protein sequence of the OB domains. While the molecular details of ssDNA binding by hSSB1 has been studied extensively, comparatively little is known about hSSB2. In this study, we use NMR solution-state backbone resonance assignments of the OB domain of hSSB2 to map the ssDNA interaction interface. Our data reveal that ssDNA binding by hSSB2 is driven by four key aromatic residues in analogy to hSSB1, however, some significant differences in the chemical shift perturbations are observed, reflecting differences in ssDNA recognition. Future studies will aim at determining the structural basis of these differences and thus help to gain a more comprehensive understanding of the functional divergences that these novel hSSBs display in the context of genome maintenance.

Keywords

DNA repair NMR hSSB1 hSSB2 SSBs 

Notes

Acknowledgements

We would like to thank Dr Ann Kwan from the University of Sydney for expert advice and maintenance of NMR spectrometers. This work was supported by an NHMRC Project Grant (1066550).

References

  1. Ashton NW, Bolderson E, Cubeddu L, O’Byrne KJ, Richard DJ (2013) Human single-stranded DNA binding proteins are essential for maintaining genomic stability. BMC Mol Biol 14:9. doi: 10.1186/1471-2199-14-9 CrossRefGoogle Scholar
  2. Ashton NW, Loo D, Paquet N, O’Byrne KJ, Richard DJ (2016) Novel insight into the composition of human single-stranded DNA-binding protein 1 (hSSB1)-containing protein complexes. BMC Mol Biol 17:24. doi: 10.1186/s12867-016-0077-5 CrossRefGoogle Scholar
  3. Ashton NW et al (2017) hSSB1 phosphorylation is dynamically regulated by DNA-PK and PPP-family protein phosphatases. DNA Repair 54:30–39. doi: 10.1016/j.dnarep.2017.03.006 CrossRefGoogle Scholar
  4. Ayed A, Mulder FA, Yi GS, Lu Y, Kay LE, Arrowsmith CH (2001) Latent and active p53 are identical in conformation. Nat Struct Biol 8:756–760. doi: 10.1038/nsb0901-756 CrossRefGoogle Scholar
  5. Bolderson E et al (2014) Human single-stranded DNA binding protein 1 (hSSB1/NABP2) is required for the stability and repair of stalled replication forks. Nucleic Acids Res 42:6326–6336. doi: 10.1093/nar/gku276 CrossRefGoogle Scholar
  6. Boucher D, Vu T, Bain AL, Tagliaro-Jahns M, Shi W, Lane SW, Khanna KK (2015) Ssb2/Nabp1 is dispensable for thymic maturation, male fertility, and DNA repair in mice. FASEB J 29:3326–3334. doi: 10.1096/fj.14-269944 CrossRefGoogle Scholar
  7. Croft LV, Ashton NW, Paquet N, Bolderson E, O’Byrne KJ, Richard DJ (2017) hSSB1 associates with and promotes stability of the BLM helicase. BMC Mol Biol 18:13. doi: 10.1186/s12867-017-0090-3 CrossRefGoogle Scholar
  8. De Braekeleer E, Douet-Guilbert N, De Braekeleer M (2014) RARA fusion genes in acute promyelocytic leukemia: a review. Expert Rev Hematol 7:347–357. doi: 10.1586/17474086.2014.903794 CrossRefGoogle Scholar
  9. Feldhahn N et al (2012) The hSSB1 orthologue Obfc2b is essential for skeletogenesis but dispensable for the DNA damage response in vivo. EMBO J 31:4045–4056. doi: 10.1038/emboj.2012.247 CrossRefGoogle Scholar
  10. Gamsjaeger R et al (2013) A structural analysis of DNA binding by myelin transcription factor 1 double zinc fingers. J Biol Chem 288:35180–35191. doi: 10.1074/jbc.M113.482075 CrossRefGoogle Scholar
  11. Gamsjaeger R, Kariawasam R, Touma C, Kwan AH, White MF, Cubeddu L (2014) Backbone and side-chain (1)H, (1)(3)C and (1)(5)N resonance assignments of the OB domain of the single stranded DNA binding protein from Sulfolobus solfataricus and chemical shift mapping of the DNA-binding interface. Biomol NMR Assign 8:243–246. doi: 10.1007/s12104-013-9492-4 CrossRefGoogle Scholar
  12. Gu P, Deng W, Lei M, Chang S (2013) Single strand DNA binding proteins 1 and 2 protect newly replicated telomeres. Cell Res 23:705–719. doi: 10.1038/cr.2013.31 CrossRefGoogle Scholar
  13. Huang J, Gong Z, Ghosal G, Chen J (2009) SOSS complexes participate in the maintenance of genomic stability. Mol Cell 35:384–393. doi: 10.1016/j.molcel.2009.06.011 CrossRefGoogle Scholar
  14. Iftode C, Daniely Y, Borowiec JA (1999) Replication protein A (RPA): the eukaryotic SSB. Crit Rev Biochem Mol Biol 34:141–180. doi: 10.1080/10409239991209255 CrossRefGoogle Scholar
  15. Kang HS, Beak JY, Kim YS, Petrovich RM, Collins JB, Grissom SF, Jetten AM (2006) NABP1, a novel RORγ-regulated gene encoding a single-stranded nucleic-acid-binding protein. Biochem J 397:89–99. doi: 10.1042/BJ20051781 CrossRefGoogle Scholar
  16. Kariawasam R, Touma C, Cubeddu L, Gamsjaeger R (2016) Backbone (1)H, (13)C and (15)N resonance assignments of the OB domain of the single stranded DNA-binding protein hSSB1 (NABP2/OBFC2B) and chemical shift mapping of the DNA-binding interface. Biomol NMR Assign 10:297–300. doi: 10.1007/s12104-016-9687-6 CrossRefGoogle Scholar
  17. Li YJ et al (2009) hSSB1 and hSSB2 form similar multiprotein complexes that participate in DNA damage response. J Biol Chem 284:23525–23531. doi: 10.1074/jbc.C109.039586 CrossRefGoogle Scholar
  18. Murzin AG (1993) OB (oligonucleotide/oligosaccharide binding)-fold: common structural and functional solution for non-homologous sequences. EMBO J 12:861Google Scholar
  19. Mushegian AR, Koonin EV (1996) A minimal gene set for cellular life derived by comparison of complete bacterial genomes. Proc Natl Acad Sci USA 93:10268–10273. doi: 10.1073/pnas.93.19.10268 ADSCrossRefGoogle Scholar
  20. Newport JW, Lonberg N, Kowalczykowski SC, von Hippel PH (1981) Interactions of bacteriophage T4-coded gene 32 protein with nucleic acids. II. Specificity of binding to DNA and RNA. J Mol Biol 145:105–121. doi: 10.1016/0022-2836(81)90336-3 CrossRefGoogle Scholar
  21. Paquet N et al (2015) HSSB1 (NABP2/OBFC2B) is required for the repair of 8-oxo-guanine by the hOGG1-mediated base excision repair pathway. Nucleic Acids Res 43:8817–8829. doi: 10.1093/nar/gkv790 CrossRefGoogle Scholar
  22. Paquet N et al (2016) hSSB1 (NABP2/OBFC2B) is regulated by oxidative stress. Sci Rep 6:27446. doi: 10.1038/srep27446 ADSCrossRefGoogle Scholar
  23. Richard DJ, Khanna KK (2009) Single-stranded DNA binding proteins involved in genome maintenance. In: Khanna K, Shiloh Y (eds) The DNA damage response: implications on cancer formation and treatment. Springer, Dordrecht, pp 349–366. doi: 10.1007/978-90-481-2561-6_16 CrossRefGoogle Scholar
  24. Richard DJ et al (2008) Single-stranded DNA-binding protein hSSB1 is critical for genomic stability. Nature 453:677–681. doi: 10.1038/nature06883 ADSCrossRefGoogle Scholar
  25. Richard DJ et al (2011a) hSSB1 interacts directly with the MRN complex stimulating its recruitment to DNA double-strand breaks and its endo-nuclease activity. Nucleic Acids Res 39:3643–3651 doi: 10.1093/Nar/Gkq1340 CrossRefGoogle Scholar
  26. Richard DJ et al (2011b) hSSB1 rapidly binds at the sites of DNA double-strand breaks and is required for the efficient recruitment of the MRN complex. Nucleic Acids Res 39:1692–1702. doi: 10.1093/nar/gkq1098 CrossRefGoogle Scholar
  27. Shi W et al (2017) Ssb1 and Ssb2 cooperate to regulate mouse hematopoietic stem and progenitor cells by resolving replicative stress. Blood 129:2479–2492. doi: 10.1182/blood-2016-06-725093 CrossRefGoogle Scholar
  28. Suck D (1997) Common fold, common function, common origin? Nat Structural Biol 4:161–165CrossRefGoogle Scholar
  29. Sun S, Shamoo Y (2003) Biochemical characterization of interactions between DNA polymerase and single-stranded DNA-binding protein in bacteriophage RB69. J Biol Chem 278:3876–3881. doi: 10.1074/jbc.M210497200 CrossRefGoogle Scholar
  30. Touma C et al (2016) A structural analysis of DNA binding by hSSB1 (NABP2/OBFC2B) in solution. Nucleic Acids Res 44:7963–7973. doi: 10.1093/nar/gkw617 CrossRefGoogle Scholar
  31. Touma C et al (2017) A data-driven structural model of hSSB1 (NABP2/OBFC2B) self-oligomerization. Nucleic Acids Res 45:8609–8620. doi: 10.1093/nar/gkx526 CrossRefGoogle Scholar
  32. Vernin C et al (2014) HTLV-1 bZIP factor HBZ promotes cell proliferation and genetic instability by activating OncomiRs. Cancer Res 74:6082–6093. doi: 10.1158/0008-5472.CAN-13-3564 CrossRefGoogle Scholar
  33. Won D et al (2013) OBFC2A/RARA: a novel fusion gene in variant acute promyelocytic leukemia. Blood 121:1432–1435. doi: 10.1182/blood-2012-04-423129 CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2017

Authors and Affiliations

  1. 1.School of Science and HealthWestern Sydney UniversityPenrithAustralia
  2. 2.School of Life and Environmental SciencesUniversity of SydneySydneyAustralia

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