Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Letter
  • Published:

Structure of the DNA deaminase domain of the HIV-1 restriction factor APOBEC3G

Nature volume 452, pages 116–119 (2008)Cite this article

Abstract

The human APOBEC3G (apolipoprotein B messenger-RNA-editing enzyme, catalytic polypeptide-like 3G) protein is a single-strand DNA deaminase that inhibits the replication of human immunodeficiency virus-1 (HIV-1), other retroviruses and retrotransposons1,2,3,4,5,6. APOBEC3G anti-viral activity is circumvented by most retroelements, such as through degradation by HIV-1 Vif7. APOBEC3G is a member of a family of polynucleotide cytosine deaminases, several of which also target distinct physiological substrates. For instance, APOBEC1 edits APOB mRNA and AID deaminates antibody gene DNA8,9,10. Although structures of other family members exist, none of these proteins has elicited polynucleotide cytosine deaminase or anti-viral activity11,12,13,14,15,16. Here we report a solution structure of the human APOBEC3G catalytic domain. Five α-helices, including two that form the zinc-coordinating active site, are arranged over a hydrophobic platform consisting of five β-strands. NMR DNA titration experiments, computational modelling, phylogenetic conservation and Escherichia coli-based activity assays combine to suggest a DNA-binding model in which a brim of positively charged residues positions the target cytosine for catalysis. The structure of the APOBEC3G catalytic domain will help us to understand functions of other family members and interactions that occur with pathogenic proteins such as HIV-1 Vif.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Functional and biophysical properties of A3G-2K3A.
Figure 2: NMR structure of A3G-2K3A.
Figure 3: The relationship of the catalytic domain of A3G to selected family members.
Figure 4: A3G catalytic domain DNA interaction model.

Similar content being viewed by others

Accession codes

Primary accessions

Protein Data Bank

Data deposits

The atomic coordinates for A3G-2K3A have been deposited in the Protein Data Bank under the accession number 2jyw.

References

  1. Chelico, L., Pham, P., Calabrese, P. & Goodman, M. F. APOBEC3G DNA deaminase acts processively 3' → 5' on single-stranded DNA. Nat. Struct. Mol. Biol. 13, 392–399 (2006)

    Article  CAS  Google Scholar 

  2. Esnault, C. et al. APOBEC3G cytidine deaminase inhibits retrotransposition of endogenous retroviruses. Nature 433, 430–433 (2005)

    Article  ADS  CAS  Google Scholar 

  3. Harris, R. S. et al. DNA deamination mediates innate immunity to retroviral infection. Cell 113, 803–809 (2003)

    Article  CAS  Google Scholar 

  4. Mangeat, B. et al. Broad antiretroviral defence by human APOBEC3G through lethal editing of nascent reverse transcripts. Nature 424, 99–103 (2003)

    Article  ADS  CAS  Google Scholar 

  5. Sheehy, A. M., Gaddis, N. C., Choi, J. D. & Malim, M. H. Isolation of a human gene that inhibits HIV-1 infection and is suppressed by the viral Vif protein. Nature 418, 646–650 (2002)

    Article  ADS  CAS  Google Scholar 

  6. Zhang, H. et al. The cytidine deaminase CEM15 induces hypermutation in newly synthesized HIV-1 DNA. Nature 424, 94–98 (2003)

    Article  ADS  CAS  Google Scholar 

  7. Yu, X. et al. Induction of APOBEC3G ubiquitination and degradation by an HIV-1 Vif–Cul5–SCF complex. Science 302, 1056–1060 (2003)

    Article  ADS  CAS  Google Scholar 

  8. Conticello, S. G., Langlois, M. A., Yang, Z. & Neuberger, M. S. DNA deamination in immunity: AID in the context of its APOBEC relatives. Adv. Immunol. 94, 37–73 (2007)

    Article  CAS  Google Scholar 

  9. Di Noia, J. M. & Neuberger, M. S. Molecular mechanisms of antibody somatic hypermutation. Annu. Rev. Biochem. 76, 1–22 (2007)

    Article  CAS  Google Scholar 

  10. Wedekind, J. E., Dance, G. S., Sowden, M. P. & Smith, H. C. Messenger RNA editing in mammals: new members of the APOBEC family seeking roles in the family business. Trends Genet. 19, 207–216 (2003)

    Article  CAS  Google Scholar 

  11. Betts, L., Xiang, S., Short, S. A., Wolfenden, R. & Carter, C. W. Cytidine deaminase. The 2.3 Å crystal structure of an enzyme: transition-state analog complex. J. Mol. Biol. 235, 635–656 (1994)

    Article  CAS  Google Scholar 

  12. Harris, R. S., Petersen-Mahrt, S. K. & Neuberger, M. S. RNA editing enzyme APOBEC1 and some of its homologs can act as DNA mutators. Mol. Cell 10, 1247–1253 (2002)

    Article  CAS  Google Scholar 

  13. Johansson, E., Mejlhede, N., Neuhard, J. & Larsen, S. Crystal structure of the tetrameric cytidine deaminase from Bacillus subtilis at 2.0 Å resolution. Biochemistry 41, 2563–2570 (2002)

    Article  CAS  Google Scholar 

  14. Ko, T. P. et al. Crystal structure of yeast cytosine deaminase. Insights into enzyme mechanism and evolution. J. Biol. Chem. 278, 19111–19117 (2003)

    Article  CAS  Google Scholar 

  15. Prochnow, C., Bransteitter, R., Klein, M. G., Goodman, M. F. & Chen, X. S. The APOBEC-2 crystal structure and functional implications for the deaminase AID. Nature 445, 447–451 (2007)

    Article  ADS  CAS  Google Scholar 

  16. Xie, K. et al. The structure of a yeast RNA-editing deaminase provides insight into the fold and function of activation-induced deaminase and APOBEC-1. Proc. Natl Acad. Sci. USA 101, 8114–8119 (2004)

    Article  ADS  CAS  Google Scholar 

  17. Iwatani, Y., Takeuchi, H., Strebel, K. & Levin, J. G. Biochemical activities of highly purified, catalytically active human APOBEC3G: correlation with antiviral effect. J. Virol. 80, 5992–6002 (2006)

    Article  CAS  Google Scholar 

  18. Chen, K. M. et al. Extensive mutagenesis experiments corroborate a structural model for the DNA deaminase domain of APOBEC3G. FEBS Lett. 581, 4761–4766 (2007)

    Article  CAS  Google Scholar 

  19. Navarro, F. et al. Complementary function of the two catalytic domains of APOBEC3G. Virology 333, 374–386 (2005)

    Article  CAS  Google Scholar 

  20. Newman, E. N. et al. Antiviral function of APOBEC3G can be dissociated from cytidine deaminase activity. Curr. Biol. 15, 166–170 (2005)

    Article  CAS  Google Scholar 

  21. Losey, H. C., Ruthenburg, A. J. & Verdine, G. L. Crystal structure of Staphylococcus aureus tRNA adenosine deaminase TadA in complex with RNA. Nat. Struct. Mol. Biol. 13, 153–159 (2006)

    Article  CAS  Google Scholar 

  22. Huthoff, H. & Malim, M. H. Cytidine deamination and resistance to retroviral infection: towards a structural understanding of the APOBEC proteins. Virology 334, 147–153 (2005)

    Article  CAS  Google Scholar 

  23. Chung, S. J., Fromme, J. C. & Verdine, G. L. Structure of human cytidine deaminase bound to a potent inhibitor. J. Med. Chem. 48, 658–660 (2005)

    Article  CAS  Google Scholar 

  24. Zhang, K. L. et al. Model structure of human APOBEC3G. PLoS ONE 2, e378 (2007)

    Article  ADS  Google Scholar 

  25. Mariani, R. et al. Species-specific exclusion of APOBEC3G from HIV-1 virions by Vif. Cell 114, 21–31 (2003)

    Article  CAS  Google Scholar 

  26. Teh, A. H. et al. The 1.48 Å resolution crystal structure of the homotetrameric cytidine deaminase from mouse. Biochemistry 45, 7825–7833 (2006)

    Article  CAS  Google Scholar 

  27. Xiang, S., Short, S. A., Wolfenden, R. & Carter, C. W. The structure of the cytidine deaminase-product complex provides evidence for efficient proton transfer and ground-state destabilization. Biochemistry 36, 4768–4774 (1997)

    Article  CAS  Google Scholar 

  28. Devany, M., Kotharu, N. P. & Matsuo, H. Solution NMR structure of the C-terminal domain of the human protein DEK. Protein Sci. 13, 2252–2259 (2004)

    Article  CAS  Google Scholar 

  29. Matsuo, H. et al. Structure of translation factor eIF4E bound to m7GDP and interaction with 4E-binding protein. Nat. Struct. Biol. 4, 717–724 (1997)

    Article  CAS  Google Scholar 

  30. Kim, S., Cullis, D. N., Feig, L. A. & Baleja, J. D. Solution structure of the Reps1 EH domain and characterization of its binding to NPF target sequences. Biochemistry 40, 6776–6785 (2001)

    Article  CAS  Google Scholar 

  31. Philo, J. S. Improved methods for fitting sedimentation coefficient distributions derived by time-derivative techniques. Anal. Biochem. 354, 238–246 (2006)

    Article  CAS  Google Scholar 

  32. Philo, J. S. A method for directly fitting the time derivative of sedimentation velocity data and an alternative algorithm for calculating sedimentation coefficient distribution functions. Anal. Biochem. 279, 151–163 (2000)

    Article  CAS  Google Scholar 

  33. Schuck, P. On the analysis of protein self-association by sedimentation velocity analytical ultracentrifugation. Anal. Biochem. 320, 104–124 (2003)

    Article  CAS  Google Scholar 

  34. Stafford, W. F. & Sherwood, P. J. Analysis of heterologous interacting systems by sedimentation velocity: curve fitting algorithms for estimation of sedimentation coefficients, equilibrium and kinetic constants. Biophys. Chem. 108, 231–243 (2004)

    Article  CAS  Google Scholar 

  35. Matsuo, H., Kupce, E., Li, H. & Wagner, G. Increased sensitivity in HNCA and HN(CO)CA experiments by selective C beta decoupling. J. Magn. Reson. B. 113, 91–96 (1996)

    Article  CAS  Google Scholar 

  36. Ikura, M., Kay, L. E. & Bax, A. A novel approach for sequential assignment of 1H, 13C, and 15N spectra of proteins: heteronuclear triple-resonance three-dimensional NMR spectroscopy. Application to calmodulin. Biochemistry 29, 4659–4667 (1990)

    Article  CAS  Google Scholar 

  37. Kay, L., Ikura, M., Tschudin, R. & Bax, A. Three-dimensional triple-resonance NMR spectroscopy of isotopically enriched proteins. J. Magn. Reson. 89, 496–514 (1990)

    ADS  CAS  Google Scholar 

  38. Wittekind, M. & Mueller, J. HNCACB, a high-sensitivity 3D NMR experiment to correlate amide-proton and nitrogen resonances with the alpha- and beta- carbon resonances in proteins. J. Magn. Reson. B. 101, 201–205 (1993)

    Article  CAS  Google Scholar 

  39. Yamazaki, T., Lee, W., Arrowsmith, C. H., Muhandiram, D. R. & Kay, L. E. A suite of triple resonance NMR experiments for the backbone assignment of 15N, 13C, 2H labeled proteins with high sensitivity. J. Am. Chem. Soc. 116, 11655–11666 (1994)

    Article  CAS  Google Scholar 

  40. Matsuo, H., Kupce, E., Li, H. & Wagner, G. Use of selective C alpha pulses for improvement of HN(CA)CO-D and HN(COCA)NH-D experiments. J. Magn. Reson. B. 111, 194–198 (1996)

    Article  CAS  Google Scholar 

  41. Matsuo, H., Li, H. & Wagner, G. A sensitive HN(CA)CO experiment for deuterated proteins. J. Magn. Reson. B. 110, 112–115 (1996)

    Article  CAS  Google Scholar 

  42. Clubb, R. T., Thanabal, V. & Wagner, G. A constant-time three-dimensional triple-resonance pulse scheme to correlate intraresidue 1HN, 15N, and 13C′ chemical shifts in 15N-13C-labeled proteins. J. Magn. Reson. 97, 213–217 (1992)

    ADS  CAS  Google Scholar 

  43. Grzesiek, S., Anglister, J. & Bax, A. Correlation of backbone amide and aliphatic side-chain resonances in 13C/15N-enriched proteins by isotropic mixing of 13C magnetization. J. Magn. Reson. B. 101, 114–119 (1993)

    Article  CAS  Google Scholar 

  44. Clore, G. M., Bax, A., Driscoll, P. C., Wingfield, P. T. & Gronenborn, A. M. Assignment of the side-chain 1H and 13C resonances of interleukin-1 beta using double- and triple-resonance heteronuclear three-dimensional NMR spectroscopy. Biochemistry 29, 8172–8184 (1990)

    Article  CAS  Google Scholar 

  45. Zhang, O., Kay, L. E., Olivier, J. P. & Forman-Kay, J. Backbone 1H and 15N resonance assignments of the N-terminal SH3 domain of drk in folded and unfolded states using enhanced-sensitivity pulsed field gradient NMR techniques. J. Biomol. NMR 4, 845–858 (1994)

    Article  CAS  Google Scholar 

  46. Delaglio, F. et al. NMRPipe: a multidimensional spectral processing system based on UNIX pipes. J. Biomol. NMR 6, 277–293 (1995)

    Article  CAS  Google Scholar 

  47. Keller, R. Optimizing the process of nuclear magnetic resonance spectrum analysis and computer aided resonance assignment. PhD thesis, Swiss Fed. Inst. Tech. Zurich (2004)

  48. Cornilescu, G., Delaglio, F. & Bax, A. Protein backbone angle restraints from searching a database for chemical shift and sequence homology. J. Biomol. NMR 13, 289–302 (1999)

    Article  CAS  Google Scholar 

  49. Herrmann, T., Guntert, P. & Wuthrich, K. Protein NMR structure determination with automated NOE-identification in the NOESY spectra using the new software ATNOS. J. Biomol. NMR 24, 171–189 (2002)

    Article  CAS  Google Scholar 

  50. Herrmann, T., Guntert, P. & Wuthrich, K. Protein NMR structure determination with automated NOE assignment using the new software CANDID and the torsion angle dynamics algorithm DYANA. J. Mol. Biol. 319, 209–227 (2002)

    Article  CAS  Google Scholar 

  51. Brunger, A. T. et al. Crystallography & NMR system: A new software suite for macromolecular structure determination. Acta Crystallogr. D Biol. Crystallogr. 54, 905–921 (1998)

    Article  CAS  Google Scholar 

  52. Desmet, J., De Maeyer, M., Hazas, B. & Lasters, I. The dead-end elimination theorem and its use in protein side-chain positioning. Nature 356, 539–542 (1992)

    Article  ADS  CAS  Google Scholar 

  53. Goldstein, R. F. Efficient rotamer elimination applied to protein side-chains and related spin glasses. Biophys. J. 66, 1335–1340 (1994)

    Article  ADS  CAS  Google Scholar 

Download references

Acknowledgements

We thank R. LaRue, N. Martemyanova, M. Stenglein and S. Wagner for assistance, laboratory members for discussions, V. Pathak for sharing unpublished information, and J. Lipscomb and K. Walters for comments on the manuscript. Key instrumentation was provided by the University of Minnesota NMR Facility (NSF) and Supercomputing Institute, the University of Wisconsin NMRfam (NIH) and the University of Connecticut Analytical Ultracentrifugation Facility. This work was supported by grants from the National Institutes of Health (A.F., H.M. and R.S.H.), the Medica Foundation (MN Partnership for Biotechnology and Medical Genomics (H.M. and R.S.H.)), the University of Minnesota (H.M. and R.S.H.) and the Searle Scholarship Program (R.S.H.).

Author Contributions H.M. and R.S.H. conceived the experimental designs, wrote the manuscript and assisted with experimentation. K.C., E.H., P.G., Y.L., A.F. and K.S. primarily contributed to protein purification, NMR data analyses, activity assays, site-directed mutagenesis, A3G-2K3A–DNA complex modelling and immunoblotting/purification optimization experiments, respectively. All authors contributed to data analyses, figure constructions and manuscript revisions.

Author information

Author notes

  1. Kuan-Ming Chen, Elena Harjes and Phillip J. Gross: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Biochemistry, Molecular Biology and Biophysics,

    Kuan-Ming Chen, Elena Harjes, Phillip J. Gross, Yongjian Lu, Keisuke Shindo, Reuben S. Harris & Hiroshi Matsuo

  2. Institute for Molecular Virology and,,

    Kuan-Ming Chen, Elena Harjes, Phillip J. Gross, Yongjian Lu, Keisuke Shindo, Reuben S. Harris & Hiroshi Matsuo

  3. Arnold and Mabel Beckman Center for Genome Engineering, University of Minnesota, Minneapolis, Minnesota 55455, USA,

    Phillip J. Gross, Keisuke Shindo & Reuben S. Harris

  4. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, 240 Longwood Avenue, Boston, Massachusetts 02115, USA,

    Amr Fahmy

Authors
  1. Kuan-Ming Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Elena Harjes
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Phillip J. Gross
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Amr Fahmy
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Yongjian Lu
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Keisuke Shindo
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Reuben S. Harris
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Hiroshi Matsuo
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Reuben S. Harris or Hiroshi Matsuo.

Supplementary information

Supplementary Information

The file contains Supplementary Discussion with additional references, Supplementary Tables S1-S3 and Supplementary Figures S1-S6 with Legends. (PDF 3551 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Chen, KM., Harjes, E., Gross, P. et al. Structure of the DNA deaminase domain of the HIV-1 restriction factor APOBEC3G. Nature 452, 116–119 (2008). https://doi.org/10.1038/nature06638

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature06638

This article is cited by

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing