Solution structure of P22 transcriptional antitermination N peptide–box B RNA complex


We have determined the solution structure of a 15-mer boxB RNA hairpin complexed with a 20-mer basic peptide of the N protein involved in bacteriophage P22 transcriptional antitermination. Complex formation involves adaptive binding with the N peptide adopting a bent α-helical conformation that packs tightly through hydrophobic and electrostatic interactions against the major groove face of the boxB RNA hairpin, orienting the open opposite face for potential interactions with host factors and/or RNA polymerase. Four nucleotides in the boxB RNA hairpin pentaloop form a stable GNRA like tetraloop structural scaffold on complex formation, allowing the looped out fifth nucleotide to make extensive hydrophobic contacts with the bound peptide. The guanidinium group of a key arginine is hydrogen-bonded to the guanine in a loop-closing sheared G·A mismatch and to adjacent backbone phosphates. The identified intermolecular contacts account for the consequences of N peptide and boxB RNA mutations on bacteriophage transcriptional antitermination.

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  1. 1

    Greenblatt, J., Nodwell, J. R. & Mason, S. W. Transcriptional antitermination. Nature 364, 401–406 (1993).

  2. 2

    Das, A. Control of transcription termination by RNA-binding proteins. Annu. rev. Biochem. 62, 893–930 (1993).

  3. 3

    Friedman, D. I. & Court, D. L. Transcriptional antitermination: the λ paradigm updated. Mol. Microbiol. 18, 191–200 (1995).

  4. 4

    Chattopadhyay, S., Garcia-Mena, J., DeVito, J., Wolska, K. & Das, A. Bipartite function of a small RNA hairpin in transcription antitermination in bacteriophage λ. Proc. Natl. Acad. Sci. USA 92, 4061–4065 (1995).

  5. 5

    Lazinski, D., Grzadzielska, E. & Das, A. Sequence-specific recognition of RNA hairpins by bacteriophage antiterminators requires a conserved arginine-rich motif. Cell 59, 207–218 (1989).

  6. 6

    Van Gilst, M. R. & von Hippel, P. H. Assembly of the N-dependent antitermination complex of phage λ: NusA and RNA bind independently to different unfolded domains of the N protein. J. Mol. Biol. 274, 160–173 (1997).

  7. 7

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

  8. 8

    Cilley, C. D. & Williamson, J. R. Analysis of bacteriophage N protein and peptide binding to boxB RNA using polyacrylamide gel coelectrophoresis (PACE). RNA 3, 57–67 (1997).

  9. 9

    Brünger, A.T. A system for X-ray crystallogrphy and NMR (Yale University Press, New Haven,-Connecticut, 1992).

  10. 10

    Jucker, F. M., Heus, H. A., Yip, P. F., Moors, E. H. & Pardi, A. A network of heterogeneous hydrogen bonds in GNRA tetraloops. J. Mol. Biol. 264, 968–980 (1996).

  11. 11

    Cate, J. H. et al. Crystal structure of a group I ribozyme domain: principles of RNA packing. Science 273, 1678–1685 (1996).

  12. 12

    Pley, H., Flaherty, K. & McKay, D. Model for an RNA tertiary interaction from the structure of an intermolecular complex between a GAAA tetraloop and an RNA helix. Nature 372, 111–113 (1994).

  13. 13

    Saenger, W. Principles of nucleic acid structure. In Springer Advanced Texts in Chemistry (ed. Cantor, C. R.) 118 (Springer-Verlag, New York; 1984).

  14. 14

    Calnan, B. J., Tidor, B., Biancalana, S., Hudson, D. & Frankel, A. D. Arginine-mediated RNA recognition: the arginine fork. Science 252, 1167–1171 (1991).

  15. 15

    Puglisi, J. D., Tan, R., Calnan, B. J., Frankel, A. D. & Williamson, J. R. Conformation of the TAR RNA-arginine complex by NMR spectroscopy. Science 257, 76–80 (1992).

  16. 16

    Franklin, N. C. Clustered arginine residues of bacteriophage λ N protein are essential to antitermination of transcription, but their locale cannot compensate for boxB loop defects. J. Mol. Biol. 231, 343–360 (1993).

  17. 17

    Su, L. et al. RNA recognition by a bent α-helix regulates transcriptional antitermination in phage λ. Biochemistry 36, 12722–12732 (1997).

  18. 18

    Su, L. et al. An RNA enhancer in a phage transcriptional antitermination complex functions as a structural switch. Genes Dev. 11, 2214–2226 (1997).

  19. 19

    Zwahlen, C. et al. Methods for measurement of intermolecular NOEs by multinuclear NMR spectroscopy: application to a bacteriophage λ N-peptide/boxB RNA complex. J. Am. Chem. Soc. 119, 6711–6721 (1997).

  20. 20

    Puglisi, J. D., Chen, L., Blanchard, S. & Frankel, A. D. Solution structure of a bovine immunodeficiency virus Tat-TAR peptide-RNA complex. Science 270, 1200–1203 (1995).

  21. 21

    Ye, X., Kumar, R. A. & Patel, D. J. Molecular recognition in the bovine immunodeficiency virus Tat peptide-TAR RNA complex. Chem. Biol. 2, 827–840 (1995).

  22. 22

    Battiste, J. L. et al. α helix-RNA major groove recognition in an HIV-1 rev peptide-RRE RNA complex. Science 273, 1547–1551 (1996).

  23. 23

    Ye, X., Gorin, A., Ellington, A. D. & Patel, D. J. Deep penetration of an α-helix into a widened RNA major groove in the HIV-1 rev peptide-RNA aptamer complex. Nature Struct. Biol. 3, 1026–1033 (1996).

  24. 24

    Rould, M. A., Perona, J. J. & Steitz, T. A. Structural basis of anticodon loop recognition by glutaminyl-tRNA synthetase. Nature 352, 213–218 (1991).

  25. 25

    Oubridge, C., Ito, N., Evans, P. R., Teo, C. H. & Nagai, K. Crystal structure at 1.92 Å resolution of the RNA-binding domain of the U1A spliceosomal protein complexed with an RNA hairpin. Nature 372, 432–438 (1994).

  26. 26

    Convery, M. A. et al. Crystal structure of an RNA aptamer-protein complex at 2.8 Å resolution. Nature Struct. Biol. 5, 133–139 (1998).

  27. 27

    Allain, F. H. et al. Specificity of ribonucleoprotein interaction determined by RNA folding during complex formation. Nature 380, 646–650 (1996).

  28. 28

    De Guzman, R. N. et al. Structure of the HIV-1 nucleocapsid protein bound to the SL3 stem-loop recognition element of the genomic ψ-RNA packaging signal. Science, 279, 384–388 (1998).

  29. 29

    Schumacher, T. N. et al. Identification of D-peptide ligands through mirror-image phage display. Science 271, 1854–1857 (1996).

  30. 30

    Milligan, J. F., Groebe, D. R., Witherell, G. W. & Uhlenbeck, O. C. Oligoribonucleotide synthesis using T7. RNA polymerase and synthetic DNA templates. Nucleic Acids Res. 15, 8783–8798 (1987).

  31. 31

    Nikonowicz, E. P. et al. Preparation of 13C and 15N labeled RNAs for heteronuclear multi-dimensional NMR studies. Nucleic Acids Res. 20, 4507–4513 (1992).

  32. 32

    Batey, R. T., Inada, M., Kujawinski, E., Puglisi, J. D. & Williamson, J. R. Preparation of isotopically labeled ribonucleotides for multidimensional NMR spectroscopy of RNA. Nucleic Acids Res. 20, 4515–4523 (1992).

  33. 33

    Deleglio, F., Grzesiek, S., Vuister, G., Zu, G., Pfeiffer, J. & Bax, A. NMRPipe: a multidimensional spectral processing system based on UNIX pipes. J. Biomol. NMR 6, 277–293 (1995).

  34. 34

    Pardi, A. Multidimensional heteronuclear NMR experiments for structure determination of isotopically labeled RNA. Meths Enz. 261, 350–380 (1995).

  35. 35

    Varani, G., Aboul-ela, F. & Allain, F. H. T. NMR investigation of RNA structure. Prog. NMR Spect. 29, 51–127 (1996).

  36. 36

    Clore, G. M. & Gronenborn, A. M. Applications of three- and four-dimensional heteronuclear NMR spectroscopy to protein structure determination. Prog. NMR Spect. 23, 43–92 (1991).

  37. 37

    Kay, L. E. Pulsed field gradient multi-dimensional NMR methods for the study of protein structure and dynamics in solution. Prog. NMR Spect. 63, 277–299 (1995).

  38. 38

    Rao, N. S. et al. NMR pulse schemes for the sequential assignment of arginine side-chain Hε protons. J. Magn. Reson. 113, 272–276 (1996).

  39. 39

    Vuister, G. W. & Bax, A. Quantitative J correlation: a new approach for measuring homonuclear three-bond J(HNHα) coupling constants in 15N-enriched proteins. J. Am. Chem. Soc. 115, 7772–7777 (1993).

  40. 40

    Nicholls, A., Sharp, K. A. & Honig, B. H. Protein folding and association: insights from the interfacial and thermodynamic properties of hydrocarbons. Proteins 11, 281–296 (1991).

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Correspondence to Dinshaw J. Patel.

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Cai, Z., Gorin, A., Frederick, R. et al. Solution structure of P22 transcriptional antitermination N peptide–box B RNA complex. Nat Struct Mol Biol 5, 203–212 (1998) doi:10.1038/nsb0398-203

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