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Antitermination in bacteriophage Î»: The structure of the N36 peptideâboxB RNA complex
- SchÃ¤rpf, Manuela, Sticht, Heinrich, Schweimer, Kristian, Boehm, Markus, Hoffmann, Silke, RÃ¶sch, Paul
- European journal of biochemistry 2000 v.267 no.8 pp. 2397-2408
- RNA, adenines, bacteriophages, electrostatic interactions, extrusion, hydrophobicity, mutants, nuclear magnetic resonance spectroscopy, nucleotides, peptides, transcription (genetics), tyrosine
- The solution structure of a 15âmer nutRboxB RNA hairpin complexed with the 36âmer Nâterminal peptide of the N protein (N36) from bacteriophage Î» was determined by 2D and 3D homonuclear and heteronuclear magnetic resonance spectroscopy. These 36 amino acids include the arginineârich motif of the N protein involved in transcriptional antitermination of phage Î». Upon complex formation with boxB RNA, the synthetic N36 peptide binds tightly to the major groove of the boxB hairpin through hydrophobic and electrostatic interactions forming a bent Î± helix. Four nucleotides of the GAAAA pentaloop of the boxB RNA adopt a GNRAâlike tetraloop fold in the complex. The formation of a GAAA tetraloop involves a loopâclosing sheared base pair (G6âA10), base stacking of three adenines (A7, A8, and A10), and extrusion of one nucleotide (A9) from the loop, as observed previously for the complex of N(1â22) peptide and the nutLboxB RNA [Legault, P., Li, J., Mogridge, J., Kay, L.E. & Greenblatt, J. (1998) Cell93, 289â299]. Stacking of the bases is extended by the indoleâring of Trp18 which also forms hydrophobic contacts to the sideâchains of Leu24, Leu25, and Val26. Based on the structure of the complex, three mutant peptides were synthesized and investigated by CD and NMR spectroscopy in order to determine the role of particular residues for complex formation. These studies revealed very distinct aminoâacid requirements at positions 3, 4, and 8, while replacement of Trp18 with tyrosine did not result in any gross structural changes.