RESUMO
During the decoding process, tRNA selection by the ribosome is far more accurate than expected from codon-anticodon pairing. Antibiotics such as streptomycin and paromomycin have long been known to increase the error rate of translation, and many mutations that increase or lower accuracy have been characterized. Recent crystal structures show that the specific recognition of base-pairing geometry leads to a closure of the domains of the small subunit around cognate tRNA. This domain closure is likely to trigger subsequent steps in tRNA selection. Many antibiotics and mutations act by making the domain closure more or less favourable. In conjunction with recent cryoelectron microscopy structures of the ribosome, a comprehensive structural understanding of the decoding process is beginning to emerge.
Assuntos
Biossíntese de Proteínas , Ribossomos/química , Ribossomos/metabolismo , Anticódon/genética , Sequência de Bases , Guanosina Trifosfato/metabolismo , Fator Tu de Elongação de Peptídeos/metabolismo , Estrutura Terciária de Proteína , RNA de Transferência Aminoácido-Específico/genética , RNA de Transferência Aminoácido-Específico/metabolismoRESUMO
The underlying basis for the accuracy of protein synthesis has been the subject of over four decades of investigation. Recent biochemical and structural data make it possible to understand at least in outline the structural basis for tRNA selection, in which codon recognition by cognate tRNA results in the hydrolysis of GTP by EF-Tu over 75 A away. The ribosome recognizes the geometry of codon-anticodon base pairing at the first two positions but monitors the third, or wobble position, less stringently. Part of the additional binding energy of cognate tRNA is used to induce conformational changes in the ribosome that stabilize a transition state for GTP hydrolysis by EF-Tu and subsequently result in accelerated accommodation of tRNA into the peptidyl transferase center. The transition state for GTP hydrolysis is characterized, among other things, by a distorted tRNA. This picture explains a large body of data on the effect of antibiotics and mutations on translational fidelity. However, many fundamental questions remain, such as the mechanism of activation of GTP hydrolysis by EF-Tu, and the relationship between decoding and frameshifting.
Assuntos
Biossíntese de Proteínas , Antibacterianos/farmacologia , Pareamento de Bases , Microscopia Crioeletrônica , Cristalografia por Raios X , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Escherichia coli/metabolismo , Expressão Gênica , Cinética , Modelos Genéticos , Modelos Moleculares , Mutação , Biossíntese de Proteínas/efeitos dos fármacos , RNA Bacteriano/química , RNA Bacteriano/genética , RNA de Transferência/química , RNA de Transferência/genética , Ribossomos/genética , Ribossomos/metabolismo , Ribossomos/ultraestruturaRESUMO
A structural and mechanistic explanation for the selection of tRNAs by the ribosome has been elusive. Here, we report crystal structures of the 30S ribosomal subunit with codon and near-cognate tRNA anticodon stem loops bound at the decoding center and compare affinities of equivalent complexes in solution. In ribosomal interactions with near-cognate tRNA, deviation from Watson-Crick geometry results in uncompensated desolvation of hydrogen-bonding partners at the codon-anticodon minor groove. As a result, the transition to a closed form of the 30S induced by cognate tRNA is unfavorable for near-cognate tRNA unless paromomycin induces part of the rearrangement. We conclude that stabilization of a closed 30S conformation is required for tRNA selection, and thereby structurally rationalize much previous data on translational fidelity.