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A ribozyme composed of only two different nucleotides


RNA molecules are thought to have been prominent in the early history of life on Earth because of their ability both to encode genetic information and to exhibit catalytic function1. The modern genetic alphabet relies on two sets of complementary base pairs to store genetic information. However, owing to the chemical instability of cytosine, which readily deaminates to uracil2, a primitive genetic system composed of the bases A, U, G and C may have been difficult to establish. It has been suggested that the first genetic material instead contained only a single base-pairing unit3,4,5,6,7. Here we show that binary informational macromolecules, containing only two different nucleotide subunits, can act as catalysts. In vitro evolution was used to obtain ligase ribozymes composed of only 2,6-diaminopurine and uracil nucleotides, which catalyse the template-directed joining of two RNA molecules, one bearing a 5′-triphosphate and the other a 3′-hydroxyl. The active conformation of the fastest isolated ribozyme had a catalytic rate that was about 36,000-fold faster than the uncatalysed rate of reaction. This ribozyme is specific for the formation of biologically relevant 3′,5′-phosphodiester linkages.

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Figure 1: Sequence and secondary structure of ligase ribozymes containing either three or two different nucleotide subunits.
Figure 2: Time course of the cyclization reaction involving the final selected ribozyme, which contained only D and U residues.
Figure 3: Saturation plot for the bimolecular reaction involving the final selected ribozyme and an RNA substrate, both of which contained only D and U residues.


  1. 1

    Gesteland, R. F., Cech, T. R. & Atkins, J. F. (eds) The RNA World 2nd edn (Cold Spring Harbor Laboratory Press, Cold Spring Harbor, 1999)

  2. 2

    Levy, M. & Miller, S. L. The stability of the RNA bases: implications for the origin of life. Proc. Natl Acad. Sci. USA 95, 7933–7938 (1998)

    ADS  CAS  Article  Google Scholar 

  3. 3

    Rich, A. Horizons in Biochemistry (eds Kasha, M. & Pullman, B.) 103–126 (Academic, New York, 1962)

    Google Scholar 

  4. 4

    Crick, F. H. C. The origin of the genetic code. J. Mol. Biol. 38, 367–379 (1968)

    CAS  Article  Google Scholar 

  5. 5

    Orgel, L. E. Evolution of the genetic apparatus. J. Mol. Biol. 38, 381–393 (1968)

    CAS  Article  Google Scholar 

  6. 6

    Wächtershäuser, G. An all-purine precursor of nucleic acids. Proc. Natl Acad. Sci. USA 85, 1134–1135 (1988)

    ADS  Article  Google Scholar 

  7. 7

    Zubay, G. An all-purine precursor of nucleic acids. Chemtracts, 2, 439–442 (1991)

    CAS  Google Scholar 

  8. 8

    Rogers, J. & Joyce, G. F. A ribozyme that lacks cytidine. Nature 402, 323–325 (1999)

    ADS  CAS  Article  Google Scholar 

  9. 9

    Rogers, J. & Joyce, G. F. The effect of cytidine on the structure and function of an RNA ligase ribozyme. RNA 7, 395–404 (2001)

    CAS  Article  Google Scholar 

  10. 10

    Kirnos, M. D., Khudyakov, I. Y., Alexandrushkina, N. I. & Vanyushin, B. F. 2-Aminoadenine is an adenine substituting for a base in S-2L cyanophage DNA. Nature 270, 369–370 (1977)

    ADS  CAS  Article  Google Scholar 

  11. 11

    Macdonald, L. E., Zhou, Y. & McAllister, W. T. Termination and slippage by bacteriophage T7 RNA polymerase. J. Mol. Biol. 232, 1030–1047 (1993)

    CAS  Article  Google Scholar 

  12. 12

    Rohatgi, R., Bartel, D. P. & Szostak, J. W. Kinetic and mechanistic analysis of nonenzymatic, template-directed oligoribonucleotide ligation. J. Am. Chem. Soc. 118, 3332–3339 (1996)

    CAS  Article  Google Scholar 

  13. 13

    Santoro, S. W. & Joyce, G. F. A general purpose RNA-cleaving DNA enzyme. Proc. Natl Acad. Sci. USA 94, 4262–4266 (1997)

    ADS  CAS  Article  Google Scholar 

  14. 14

    Orò, J. Mechanism of synthesis of adenine from hydrogen cyanide under plausible primitive Earth conditions. Nature 191, 1193–1194 (1961)

    ADS  Article  Google Scholar 

  15. 15

    Sanchez, R. A., Ferris, J. P. & Orgel, L. E. Studies in prebiotic synthesis IV. Conversion of 4-aminoimidazole-5-carbonitrile derivatives to purines. J. Mol. Biol. 38, 121–128 (1968)

    CAS  Article  Google Scholar 

  16. 16

    Joyce, G. F. Evolutionary chemistry: getting there from here. Science 276, 1658–1659 (1997)

    CAS  Article  Google Scholar 

  17. 17

    Taylor, S. V., Walter, K. U., Kast, P. & Hilvert, D. Searching sequence space for protein catalysts. Proc. Natl Acad. Sci. USA 98, 10596–10601 (2001)

    ADS  CAS  Article  Google Scholar 

  18. 18

    Plaxco, K. W., Riddle, D. S., Grantcharova, V. & Baker, D. Simplified proteins: minimalist solutions to the ‘protein folding problem’. Curr. Opin. Struct. Biol. 8, 80–85 (1998)

    CAS  Article  Google Scholar 

  19. 19

    McGinness, K. E., Wright, M. C. & Joyce, G. F. Continuous in vitro evolution of a ribozyme that catalyzes three successive nucleotidyl addition reactions. Chem. Biol. 9, 585–596 (2002)

    CAS  Article  Google Scholar 

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We thank J. Rogers for many discussions during the initial stages of this project. We also thank the members of the Joyce laboratory for their advice and E. Tzima for assistance in preparation of the manuscript. This work was supported by a grant from the National Aeronautics and Space Administration and the Skaggs Institute for Chemical Biology. J.S.R. was supported by a postdoctoral fellowship from the NASA Specialized Center for Research and Training (NSCORT) in Exobiology.

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Correspondence to Gerald F. Joyce.

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Reader, J., Joyce, G. A ribozyme composed of only two different nucleotides. Nature 420, 841–844 (2002).

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