title: Codon-anticodon interaction at the P site is a prerequisite for tRNA interaction with the small ribosomal subunit.
abstract: The arrival of high resolution crystal structures for the ribosomal subunits opens a new phase of molecular analysis and asks for corresponding analyses of ribosomal function. Here we apply the phosphorothioate technique to dissect tRNA interactions with the ribosome. We demonstrate that a tRNA bound to the P site of non-programmed 70 S ribosomes contacts predominantly the 50 S, as opposed to the 30 S subunit, indicating that codon-anticodon interaction at the P site is a prerequisite for 30 S binding. Protection patterns of tRNAs bound to isolated subunits and programmed 70 S ribosomes were compared. The results suggest the presence of a movable domain in the large ribosomal subunit that carries tRNA and reveal that only approximately 15% of a tRNA, namely residues 30 +/- 1 to 43 +/- 1, contact the 30 S subunit of programmed 70 S ribosomes, whereas the remaining 85% make contact with the 50 S subunit. Identical protection patterns of two distinct elongator tRNAs at the P site were identified as tRNA species-independent phosphate backbone contacts. The sites of protection correlate nicely with the predicted ribosomal-tRNA contacts deduced from a 5.5-A crystal structure of a programmed 70 S ribosome, thus refining which ribosomal components are critical for tRNA fixation at the P site.
authors.affiliation: Max-Planck-Institut für Molekulare Genetik, AG Ribosomen, Ihnestrasse 73, D-14195 Berlin, Germany.
authors.forename: Markus A
authors.initials: MA
authors.lastname: Schäfer
authors.forename: A Ozlem
authors.initials: AO
authors.lastname: Tastan
authors.forename: Sebastian
authors.initials: S
authors.lastname: Patzke
authors.forename: Gregor
authors.initials: G
authors.lastname: Blaha
authors.forename: Christian M T
authors.initials: CM
authors.lastname: Spahn
authors.forename: Daniel N
authors.initials: DN
authors.lastname: Wilson
authors.forename: Knud H
authors.initials: KH
authors.lastname: Nierhaus
mesh_terms: D000926:Anticodon; D001483:Base Sequence; D003062:Codon; D017931:DNA Primers; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D012343:RNA, Transfer / Q000737:chemistry / Q000378:metabolism*; D012270:Ribosomes / Q000378:metabolism*
chemical_list: D000926:Anticodon; D003062:Codon; D017931:DNA Primers; D012343:RNA, Transfer