RNA Secondary Structure Prediction

David H. Mathews1, Douglas H. Turner1, Richard M. Watson1

1 University of Rochester, Rochester
Publication Name:  Current Protocols in Nucleic Acid Chemistry
Unit Number:  Unit 11.2
DOI:  10.1002/cpnc.19
Online Posting Date:  December, 2016
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In this unit, protocols are provided for predicting RNA secondary structure with the user‐friendly RNAstructure desktop computer program and the RNAstructure Web server. The minimum free energy structure and a set of suboptimal structures with similar free energies are predicted. Prediction of high‐affinity oligonucleotide binding sites to a structured RNA target is also presented. © 2016 by John Wiley & Sons, Inc.

Keywords: RNA secondary structure prediction; free energy minimization; thermodynamics; binding affinity; RNA folding; partition function

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Table of Contents

  • Introduction
  • Basic Protocol 1: Predicting Secondary Structure and Base‐Pair Probabilities
  • Basic Protocol 2: Predicting Binding Affinities of Oligonucleotides Complementary to an RNA Target with OligoWalk
  • Basic Protocol 3: Predicting a Secondary Structure with the RNAstructure WEB Server
  • Commentary
  • Literature Cited
  • Figures
  • Tables
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Literature Cited

  Bellaousov S. and Mathews D.H. 2010. ProbKnot: Fast prediction of RNA secondary structure including pseudoknots. RNA 16:1870‐1880. doi: 10.1261/rna.2125310.
  Bohula, E.A., Salisbury, A.J., Sohail, M., Playford, M.P., Riedemann, J., Southern, E.M., and Macaulay, V.M. 2003. The efficacy of small interfering RNAs targeted to the type 1 insulin‐like growth factor receptor (IGF1R) is influenced by secondary structure in the IGF1R transcript. J. Biol. Chem. 278:15991‐15997. doi: 10.1074/jbc.M300714200.
  Brown, J.W. 1999. The ribonuclease P database. Nucleic Acids Res. 27:314. doi: 10.1093/nar/27.1.314.
  Burgstaller, P. and Famulok, M. 1997. Flavin‐dependent photocleavage of RNA at G.U base pairs. J. Am. Chem. Soc. 119:1137‐1138. doi: 10.1021/ja962918p.
  Deigan K.E., Li T.W., Mathews D.H., and Weeks K.M. 2009. Accurate SHAPE‐directed RNA structure determination. Proc. Natl. Acad. Sci. U.S.A. 106:97‐102. doi: 10.1073/pnas.0806929106.
  Dirks, R. and Pierce, N. 2003. A partition function algorithm for nucleic acid secondary structure including pseudoknots. J. Comput. Chem. 24:1664‐1677. doi: 10.1002/jcc.10296.
  Dowell, R.D. and Eddy, S.R. 2004. Evaluation of several lightweight stochastic context‐free grammars for RNA secondary structure prediction. BMC Bioinformatics 5:71. doi: 10.1186/1471‐2105‐5‐71.
  Eddy, S.R. 2004. How do RNA folding algorithms work? Nat. Biotechnol. 22:1457‐1458. doi: 10.1038/nbt1104‐1457.
  Ehresmann, C., Baudin, F., Mougel, M., Romby, P., Ebel, J., and Ehresmann, B. 1987. Probing the structure of RNAs in solution. Nucleic Acids Res. 15:9109‐9128. doi: 10.1093/nar/15.22.9109.
  Far, R.K. and Sczakiel, G. 2003. The activity of siRNA in mammalian cells is related to structural target accessibility: A comparison with antisense oligonucleotides. Nucleic Acids Res. 31:4417‐4424. doi: 10.1093/nar/gkg649.
  Heale, B.S., Soifer, H.S., Bowers, C., and Rossi, J.J. 2005. siRNA target site secondary structure predictions using local stable substructures. Nucleic Acids Res. 33:e30. doi: 10.1093/nar/gni026.
  Hofacker, I.L. 2003. Vienna RNA secondary structure server. Nucleic Acids Res. 31:3429‐3431. doi: 10.1093/nar/gkg599.
  Knapp, G. 1989. Enzymatic approaches to probing RNA secondary and tertiary structure. Methods Enzymol. 180:192‐212. doi: 10.1016/0076‐6879(89)80102‐8.
  Lu Z.J. and Mathews D.H. 2007. Efficient siRNA selection using hybridization thermodynamics. Nucleic Acids Res. 36:640‐647. doi: 10.1093/nar/gkm920.
  Lu, Z.J., Turner, D.H., and Mathews, D.H. 2006. A set of nearest neighbor parameters for predicting the enthalpy change of RNA secondary structure formation. Nucleic Acids Res. 34:4912‐4924. doi: 10.1093/nar/gkl472.
  Lu Z.J., Gloor J.W., and Mathews D.H. 2009. Improved RNA secondary structure prediction by maximizing expected pair accuracy. RNA 15:1805‐1813. doi: 10.1261/rna.1643609.
  Mathews, D.H. 2004. Using an RNA secondary structure partition function to determine confidence in base pairs predicted by free energy minimization. RNA 10:1178‐1190. doi: 10.1261/rna.7650904.
  Mathews, D.H. and Zuker, M. 2004. Predictive methods using RNA sequences. In Bioinformatics: A Practical Guide to the Analysis of Genes and Proteins, 3rd ed. (A. Baxevenis and F. Oullette, eds.) pp. 143‐170. John Wiley & Sons, Hoboken, N.J.
  Mathews, D.H., Sabina, J., Zuker, M., and Turner, D.H. 1999a. Expanded sequence dependence of thermodynamic parameters provides improved prediction of RNA secondary structure. J. Mol. Biol. 288:911‐940. doi: 10.1006/jmbi.1999.2700.
  Mathews, D.H., Burkard, M.E., Freier, S.M., Wyatt, J.R., and Turner, D.H. 1999b. Predicting oligonucleotide affinity to nucleic acid targets. RNA 5:1458‐1469. doi: 10.1017/S1355838299991148.
  Mathews, D.H., Disney, M.D., Childs, J.L., Schroeder, S.J., Zuker, M., and Turner, D.H. 2004. Incorporating chemical modification constraints into a dynamic programming algorithm for prediction of RNA secondary structure. Proc. Natl. Acad. Sci. U.S.A. 101:7287‐7292. doi: 10.1073/pnas.0401799101.
  Matveeva, O.V., Mathews, D.H., Tsodikov, A.D., Shabalina, S.A., Gesteland, R.F., Atkins, J.F., and Freier, S.M. 2003. Thermodynamic criteria for high hit rate antisense oligonucleotide design. Nucleic Acids Res. 31:4989‐4994. doi: 10.1093/nar/gkg710.
  Petch, A.K., Sohail, M., Hughes, M.D., Benter, I., Darling, J., Southern, E.M., and Akhtar, S. 2003. Messenger RNA expression profiling of genes involved in epidermal growth factor receptor signalling in human cancer cells treated with scanning array‐designed antisense oligonucleotides. Biochem. Pharmacol. 66:819‐830. doi: 10.1016/S0006‐2952(03)00407‐6.
  Rivas, E. and Eddy, S.R. 1999. A dynamic programming algorithm for RNA structure prediction including pseudoknots. J. Mol. Biol. 285:2053‐2068. doi: 10.1006/jmbi.1998.2436.
  Sprinzl M. and Vassilenko K.S. 2005. Compilation of tRNA sequences and sequences of tRNA genes. Nucleic Acids Res. 33:D139‐140. doi: 10.1093/nar/gki012.
  Williams, K.P. and Bartel, D.P. 1996. Phylogenetic analysis of tmRNA secondary structure. RNA 2:1306‐1310.
  Xia, T., SantaLucia, J. Jr., Burkard, M.E., Kierzek, R., Schroeder, S.J., Jiao, X., Cox, C., and Turner, D.H. 1998. Thermodynamic parameters for an expanded nearest‐neighbor model for formation of RNA duplexes with Watson‐Crick pairs. Biochemistry 37:14719‐14735. doi: 10.1021/bi9809425.
  Ziehler, W.A. and Engelke, D.R. 2000. Probing RNA structure with chemical reagents and enzymes. Curr. Protoc. Nucleic Acid Chem. 00:6.1.1‐6.1.21.
  Zuker, M. 1989. On finding all suboptimal foldings of an RNA molecule. Science 244:48‐52. doi: 10.1126/science.2468181.
  Zuker, M. and Jacobson, A.B. 1995. “Well‐determined” regions in RNA secondary structure predictions. Applications to small and large subunit rRNA. Nucleic Acids Res. 23:2791‐2798. doi: 10.1093/nar/23.14.2791.
Key References
  Mathews et al. 1999b, 2004. See above.
  These publications derive the thermodynamic parameters used by the secondary structure prediction algorithm and tabulate the accuracy of the algorithm with a large database of structures from sequence comparisons.
  Zuker, 1989. See above.
  Explains the method for predicting suboptimal structures using a dynamic programming algorithm.
Internet Resources
  The Mathews Lab homepage is the source for downloading RNAstructure and for running the RNAstructure Web servers.
  The XRNA homepage at the Santa Cruz RNA Center is the source of XRNA, which can be used for creating publication‐quality RNA secondary structure diagrams.
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