Principles of stop-codon reading on the ribosome.
In termination of protein synthesis, the bacterial release factors RF1 and RF2 bind to the ribosome through specific recognition of messenger RNA stop codons and trigger hydrolysis of the bond between the nascent polypeptide and the transfer RNA at the peptidyl-tRNA site, thereby releasing the newly synthesized protein. The release factors are highly specific for a U in the first stop-codon position and recognize different combinations of purines in the second and third positions, with RF1 reading UAA and UAG and RF2 reading UAA and UGA. With recently determined crystal structures of termination complexes, it has become possible to decipher the energetics of stop-codon reading by computational analysis and to clarify the origin of the high release-factor binding accuracy. Here we report molecular dynamics free-energy calculations on different cognate and non-cognate termination complexes. The simulations quantitatively explain the basic principles of decoding in all three codon positions and reveal the key elements responsible for specificity of the release factors. The overall reading mechanism involves hitherto unidentified interactions and recognition switches that cannot be described in terms of a tripeptide anticodon model. Further simulations of complexes with tRNA(Trp), the tRNA recognizing the triplet codon for Trp, explain the observation of a 'leaky' stop codon and highlight the fundamentally different third position reading by RF2, which leads to a high stop-codon specificity with strong discrimination against the Trp codon. The simulations clearly illustrate the versatility of codon reading by protein, which goes far beyond tRNA mimicry.
Department of Cell and Molecular Biology, Uppsala University, Biomedical Center, Box 596, SE-751 24 Uppsala, Sweden.
Johan
J
Sund
Martin
M
Andér
Johan
J
Aqvist
D001419:Bacteria / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018388:Codon, Terminator / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D010454:Peptide Termination Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding / Q000235:genetics; D017434:Protein Structure, Tertiary; D012343:RNA, Transfer / Q000378:metabolism; D012270:Ribosomes / Q000235:genetics / Q000378:metabolism*
D018388:Codon, Terminator; D010454:Peptide Termination Factors; D012343:RNA, Transfer