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.

  • Article
  • Published:

Oligoarginine peptides slow strand annealing and assist non-enzymatic RNA replication

A Retraction to this article was published on 23 November 2017

This article has been updated

Abstract

The non-enzymatic replication of RNA is thought to have been a critical process required for the origin of life. One unsolved difficulty with non-enzymatic RNA replication is that template-directed copying of RNA results in a double-stranded product. After strand separation, rapid strand reannealing outcompetes slow non-enzymatic template copying, which renders multiple rounds of RNA replication impossible. Here we show that oligoarginine peptides slow the annealing of complementary oligoribonucleotides by up to several thousand-fold; however, short primers and activated monomers can still bind to template strands, and template-directed primer extension can still occur, all within a phase-separated condensed state, or coacervate. Furthermore, we show that within this phase, partial template copying occurs even in the presence of full-length complementary strands. This method to enable further rounds of replication suggests one mechanism by which short non-coded peptides could have enhanced early cellular fitness, and potentially explains how longer coded peptides, that is, proteins, came to prominence in modern biology.

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

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

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

Figure 1: The reannealing problem and a proposed solution.
Figure 2: RNA–peptide binding measured by fluorescence anisotropy and circular dichroism (CD).
Figure 3: RNA-annealing rates in the presence of peptides.
Figure 4: Non-enzymatic RNA polymerization.
Figure 5: Condensed phase of RNA.

Similar content being viewed by others

Change history

  • 12 October 2017

    We the authors are retracting this Article because our efforts to repeat and follow up on the results have been unsuccessful. Specifically, we have been unable to reproduce observations suggesting that arginine-rich peptides allow the non-enzymatic copying of an RNA template in the presence of its complementary strand (Fig. 4e). We originally dismissed variability in these experiments as resulting from variability in the snap cooling of samples following thermal denaturation. However, we now understand that the data reported in the published article are the result of false positives that arose from an incorrectly designed experiment in which random errors, including transfer and concentration errors, affected the ratio of the concentrations of the RNA template and its complementary strand. This resulted in false positives that were misinterpreted as template copying in the presence of a complementary strand, where in reality these reactions did not contain enough complementary strands to completely inhibit the reaction. Subsequent experiments suggested that arginine-rich peptides may not slow the reannealing of complementary strands (Fig. 3), and that what we had previously interpreted as a decrease in annealing rate was actually an artefact due to slow coalescence or strand exchange between droplets of RNA–peptide coacervate, as well as droplet coalescence and settling that led to decreased fluorescence intensity. Similarly, the changing circular dichroism spectra shown in Figure 2c, which were originally interpreted to be the result of a change in the global helical structure of RNA upon peptide binding, may also be an artefact due to, for example, loss of signal or light scattering. Although the binding of arginine-rich peptides to RNA does form condensed-phase droplets, and although most of the RNA does reside within the condensed phase, follow-up experiments to confirm that non-enzymatic RNA polymerization occurs within these coacervate droplets have been inconclusive (Fig. 5d). The experiments showing that vesicles are stable in the presence of arginine-rich peptides (Supplementary Figure 26, by N. Kamat), and the failure of acidic peptides to condense RNA (Supplementary Figure 8, by K. Adamala) have been reproduced. However, since the main conclusions of our paper are incorrect, all of the authors are now retracting the Article. The authors would like to thank Dr Tivoli Olsen for her extensive efforts to unravel the errors in our Article and we apologize to the scientific community for any confusion arising from our publication.

References

  1. Orgel, L. E. Some consequences of the RNA world hypothesis. Orig. Life Evol. Biosph. 33, 211–218 (2003).

    CAS  PubMed  Google Scholar 

  2. Gilbert, W. Origin of life: the RNA world. Nature 319, 618 (1986).

    Article  Google Scholar 

  3. Robertson, M. P. & Joyce, G. F. The origins of the RNA world. Cold Spring Harb. Perspect. Biol. 4, a003608 (2012).

    PubMed  PubMed Central  Google Scholar 

  4. Powner, M. W., Gerland, B. & Sutherland, J. D. Synthesis of activated pyrimidine ribonucleotides in prebiotically plausible conditions. Nature 459, 239–242 (2009).

    CAS  PubMed  Google Scholar 

  5. Powner, M. W., Sutherland, J. D. & Szostak, J. W. Chemoselective multicomponent one-pot assembly of purine precursors in water. J. Am. Chem. Soc. 132, 16677–16688 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  6. Bowler, F. R. et al. Prebiotically plausible oligoribonucleotide ligation facilitated by chemoselective acetylation. Nature Chem. 5, 383–389 (2013).

    CAS  Google Scholar 

  7. Szostak, J. W. The eightfold path to non-enzymatic RNA replication. J. Syst. Chem. 3, 2 (2012).

    CAS  Google Scholar 

  8. Heuberger, B. D., Pal, A., Del Frate, F., Topkar, V. V. & Szostak, J. W. Replacing uridine with 2-thiouridine enhances the rate and fidelity of nonenzymatic RNA primer extension. J. Am. Chem. Soc. 137, 2769–2775 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  9. Adamala, K. & Szostak, J. W. Nonenzymatic template-directed RNA synthesis inside model protocells. Science 342, 1098–1100 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  10. Blain, J. C. & Szostak, J. W. Progress toward synthetic cells. Annu. Rev. Biochem. 83, 615–640 (2014).

    CAS  PubMed  Google Scholar 

  11. Deck, C., Jauker, M. & Richert, C. Efficient enzyme-free copying of all four nucleobases templated by immobilized RNA. Nature Chem. 3, 603–608 (2011).

    CAS  Google Scholar 

  12. Engelhart, A. E., Powner, M. W. & Szostak, J. W. Functional RNAs exhibit tolerance for non-heritable 2′–5′ versus 3′–5′ backbone heterogeneity. Nature Chem. 5, 390–394 (2013).

    CAS  Google Scholar 

  13. Ross, P. D. & Sturtevant, J. M. The kinetics of double helix formation from polyriboadenylic acid and polyribouridylic acid. Proc. Natl Acad. Sci. USA 46, 1360–1365 (1960).

    CAS  PubMed  Google Scholar 

  14. Izgu, E. C. et al. Uncovering the thermodynamics of monomer binding for RNA replication. J. Am. Chem. Soc. 137, 6373–6382 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  15. Kunin, V. A system of two polymerases—a model for the origin of life. Orig. Life Evol. Biosph. 30, 459–466 (2000).

    CAS  PubMed  Google Scholar 

  16. Ritson, D. J. & Sutherland, J. D. Synthesis of aldehydic ribonucleotide and amino acid precursors by photoredox chemistry. Angew. Chem. Int. Ed. 52, 5845–5847 (2013).

    CAS  Google Scholar 

  17. Mullen, L. B. & Sutherland, J. D. Simultaneous nucleotide activation and synthesis of amino acid amides by a potentially prebiotic multi-component reaction. Angew. Chem. Int. Ed. 46, 8063–8066 (2007).

    CAS  Google Scholar 

  18. Miller, S. L. A production of amino acids under possible primitive Earth conditions. Science 117, 528–529 (1953).

    CAS  Google Scholar 

  19. Plankensteiner, K., Reiner, H. & Rode, B. M. Amino acids on the rampant primordial Earth: electric discharges and the hot salty ocean. Mol. Divers. 10, 3–7 (2006).

    CAS  PubMed  Google Scholar 

  20. Ehrenfreund, P. & Cami, J. Cosmic carbon chemistry: from the interstellar medium to the early Earth. Cold Spring Harb. Perspect. Biol. 2, a002097 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  21. Pizzarello, S. & Shock, E. The organic composition of carbonaceous meteorites: the evolutionary story ahead of biochemistry. Cold Spring Harb. Perspect. Biol. 2, a002105 (2010).

    PubMed  PubMed Central  Google Scholar 

  22. Miller, S. L. Which organic compounds could have occurred on the prebiotic earth? Cold Spring Harb. Symp. Quant. Biol. 52, 17–27 (1987).

    CAS  PubMed  Google Scholar 

  23. Zaia, D. A. M., Zaia, C. T. B. V. & De Santana, H. Which amino acids should be used in prebiotic chemistry studies? Orig. Life Evol. Biosph. 38, 469–488 (2008).

    CAS  PubMed  Google Scholar 

  24. Patel, B. H., Percivalle, C., Ritson, D. J., Duffy, C. D. & Sutherland, J. D. Common origins of RNA, protein and lipid precursors in a cyanosulfidic protometabolism. Nature Chem. 7, 301–307 (2015).

    CAS  Google Scholar 

  25. Mascotti, D. P. & Lohman, T. M. Thermodynamics of oligoarginines binding to RNA and DNA. Biochemistry 36, 7272–7279 (1997).

    CAS  PubMed  Google Scholar 

  26. Tan, R. & Frankel, A. D. Structural variety of arginine-rich RNA-binding peptides. Proc. Natl Acad. Sci. USA 92, 5282–5286 (1995).

    CAS  PubMed  Google Scholar 

  27. Balhorn, R., Brewer, L. & Corzett, M. DNA condensation by protamine and arginine-rich peptides: analysis of toroid stability using single DNA molecules. Mol. Reprod. Dev. 56, 230–234 (2000).

    CAS  PubMed  Google Scholar 

  28. Hud, N. V., Allen, M. J., Downing, K. H., Lee, J. & Balhorn, R. Identification of the elemental packing unit of DNA in mammalian sperm cells by atomic force microscopy. Biochem. Biophys. Res. Commun. 193, 1347–1354 (1993).

    CAS  PubMed  Google Scholar 

  29. DeRouchey, J., Hoover, B. & Rau, D. C. A comparison of DNA compaction by arginine and lysine peptides: a physical basis for arginine rich protamines. Biochemistry 52, 3000–3009 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  30. Danger, G., Plasson, R. & Pascal, R. Pathways for the formation and evolution of peptides in prebiotic environments. Chem. Soc. Rev. 41, 5416–5429 (2012).

    CAS  PubMed  Google Scholar 

  31. Imai, E.-I., Honda, H., Hatori, K., Brack, A. & Matsuno, K. Elongation of oligopeptides in a simulated submarine hydrothermal system. Science 283, 831–833 (1999).

    CAS  PubMed  Google Scholar 

  32. Lambert, J.-F. Adsorption and polymerization of amino acids on mineral surfaces: a review. Orig. Life Evol. Biosph. 38, 211–242 (2008).

    CAS  PubMed  Google Scholar 

  33. Leman, L., Orgel, L. & Ghadiri, M. R. Carbonyl sulfide-mediated prebiotic formation of peptides. Science 306, 283–286 (2004).

    CAS  Google Scholar 

  34. Leman, L. J., Huang, Z.-Z. & Ghadiri, M. R. Peptide bond formation in water mediated by carbon disulfide. Astrobiology 15, 709–716 (2015).

    CAS  PubMed  Google Scholar 

  35. Xin, L. et al. 6-Membered ring intermediates in polymerization of N-carboxyanhydride-L-α-arginine in H2O. Sci. China Ser. B Chem. 52, 1220–1226 (2009).

    CAS  Google Scholar 

  36. Liu, R. & Orgel, L. E. Polymerization on the rocks: β-amino acids and arginine. Orig. Life Evol. Biosph. 28, 245–257 (1998).

    CAS  PubMed  Google Scholar 

  37. Hall, K. B. 2-Aminopurine as a probe of RNA conformational transitions. Methods Enzymol. 469, 269–285 (2009).

    CAS  PubMed  Google Scholar 

  38. Liu, G. et al. Biological properties of poly-L-lysine-DNA complexes generated by cooperative binding of the polycation. J. Biol. Chem. 276, 34379–34387 (2001).

    CAS  PubMed  Google Scholar 

  39. Fasting, C. et al. Multivalency as a chemical organization and action principle. Angew. Chem. Int. Ed. 51, 10472–10498 (2012).

    CAS  Google Scholar 

  40. Kypr, J., Kejnovská, I., Renčiuk, D. & Vorlíčková, M. Circular dichroism and conformational polymorphism of DNA. Nucleic Acids Res. 37, 1713–1725 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  41. Jia, T. Z., Hentrich, C. & Szostak, J. W. Rapid RNA exchange in aqueous two-phase system and coacervate droplets. Orig. Life Evol. Biosph. 44, 1–12 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  42. Frankel, E. A., Bevilacqua, P. C. & Keating, C. D. Polyamine/nucleotide coacervates provide strong compartmentalization of Mg2+, nucleotides, and RNA. Langmuir 32, 2041–2049 (2016).

    CAS  PubMed  Google Scholar 

  43. Kashiwagi, N., Furuta, H. & Ikawa, Y. Primitive templated catalysis of a peptide ligation by self-folding RNAs. Nucleic Acids Res. 37, 2574–2583 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  44. Rajkowitsch, L. et al. RNA chaperones, RNA annealers and RNA helicases. RNA Biol. 4, 118–130 (2007).

    CAS  PubMed  Google Scholar 

  45. Ziebarth, J. & Wang, Y. Molecular dynamics simulations of DNA-polycation complex formation. Biophys. J. 97, 1971–1983 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  46. Baeza, I. et al. Possible prebiotic significance of polyamines in the condensation, protection, encapsulation, and biological properties of DNA. Orig. Life Evol. Biosph. 21, 225–242 (1992).

    CAS  Google Scholar 

  47. Frederix, P. W. J. M. et al. Exploring the sequence space for (tri-)peptide self-assembly to design and discover new hydrogels. Nature Chem. 7, 30–37 (2015).

    CAS  Google Scholar 

  48. Cisse, I. I., Kim, H. & Ha, T. A rule of seven in Watson–Crick base-pairing of mismatched sequences. Nature Struct. Mol. Biol. 19, 623–627 (2012).

    CAS  Google Scholar 

  49. Pörschke, D. & Eigen, M. Co-operative non-enzymatic base recognition III. Kinetics of the helix–coil transition of the oligoribouridylic · oligoriboadenylic acid system and of oligoriboadenylic acid alone at acidic pH. J. Mol. Biol. 62, 361–381 (1971).

    PubMed  Google Scholar 

  50. Kreysing, M., Keil, L., Lanzmich, S. & Braun, D. Heat flux across an open pore enables the continuous replication and selection of oligonucleotides towards increasing length. Nature Chem. 7, 203–208 (2015).

    CAS  Google Scholar 

  51. Hurwitz, S., Harris, R. N., Werner, C. A. & Murphy, F. Heat flow in vapor dominated areas of the Yellowstone Plateau Volcanic Field: implications for the thermal budget of the Yellowstone Caldera. J. Geophys. Res. Solid Earth 117, B10207 (2012).

    Google Scholar 

  52. Dora Tang, T.-Y. et al. Fatty acid membrane assembly on coacervate microdroplets as a step towards a hybrid protocell model. Nature Chem. 6, 527–533 (2014).

    CAS  Google Scholar 

  53. Kamat, N. P., Tobé, S., Hill, I. T. & Szostak, J. W. Electrostatic localization of RNA to protocell membranes by cationic hydrophobic peptides. Angew. Chem. Int. Ed. 54, 11735–11739 (2015).

    CAS  Google Scholar 

  54. Aumiller, W. M. Jr & Keating, C. D. Phosphorylation-mediated RNA/peptide complex coacervation as a model for intracellular liquid organelles. Nature Chem. 8, 129–137 (2016).

    CAS  Google Scholar 

  55. Lin, Y., Protter, D. S. W., Rosen, M. K. & Parker, R. Formation and maturation of phase-separated liquid droplets by RNA-binding proteins. Mol. Cell 60, 208–219 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  56. Brangwynne, C. P. et al. Germline P granules are liquid droplets that localize by controlled dissolution/condensation. Science 324, 1729–1732 (2009).

    CAS  Google Scholar 

  57. Hyman, A. A. & Brangwynne, C. P. Beyond stereospecificity: liquids and mesoscale organization of cytoplasm. Dev. Cell 21, 14–16 (2011).

    CAS  PubMed  Google Scholar 

  58. Joyce, G. F., Inoue, T. & Orgel, L. E. Non-enzymatic template-directed synthesis on RNA random copolymers. Poly(C, U) templates. J. Mol. Biol. 176, 279–306 (1984).

    CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We thank A. E. Engelhart, C. Hentrich, B. D. Heuberger, A. T. Larsen, T. J. Olsen, N. Prywes, R. Saganty, L. Zhou and all other members of the Szostak Lab for helpful discussions and critical reading of the manuscript. We also thank G. Ruvkun and E. Rubin for their support and very helpful advice. J.W.S. is an Investigator of the Howard Hughes Medical Institute. A.C.F. is supported by a Research Fellowship from the Earth-Life Science Institute at the Tokyo Institute of Technology. N.P.K. is supported by an appointment to the National Aeronautics and Space Administration (NASA) Postdoctoral Program, administered by Oak Ridge Associated Universities through a contract with NASA. This work was supported by grants from the Simons Foundation (290363) and NASA (NNX15AL18G) to J.W.S.

Author information

Authors and Affiliations

Authors

Contributions

T.Z.J., A.C.F. and J.W.S. conceived the experiments and wrote the manuscript. N.P.K. performed the vesicle-leakage assays and T.Z.J. performed all the other experiments. K.P.A. contributed intellectually.

Corresponding author

Correspondence to Jack W. Szostak.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary information

Supplementary information (PDF 3204 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Jia, T., Fahrenbach, A., Kamat, N. et al. Oligoarginine peptides slow strand annealing and assist non-enzymatic RNA replication. Nature Chem 8, 915–921 (2016). https://doi.org/10.1038/nchem.2551

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nchem.2551

This article is cited by

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