We develop a quantitative theory of kinetic proofreading with an arbitrary number of checking steps after the hydrolysis of a nucleoside triphosphate. In particular, we investigate the relationship between the minimum dissipation of free energy required for a given error frequency in such systems. Several conclusions can be drawn from the present treatment: first, the ultimate accuracy of error correcting selective pathways is set by the displacement from equilibrium of the nucleoside triphosphates. Second, it is advantageous to achieve a desired accuracy at a small energy dissipation with several checking steps rather than a single one. This could explain antinomies in the amino acylation reaction as well as in mRNA translation, where small structural differences lead to large differences in flow rates between right and wrong substrates. Third, all checking steps should contribute equally to the accuracy, which implies a specific and symmetrical set of rate constants for the checking events on the enzyme.
We suggest that the interaction between a codon and its cognate tRNA induces conformational changes in the tRNA. We further suggest that sites on the ribosome preferentially bind tRNA in those conformations which require proper matching of codon and anticodon. According to this model, the codon functions as an allosteric effector which influences the conformation at various sites in the tRNA. This is made possible by the ribosome, which we suggest traps tRNA molecules in those conformation states that maximize the energy difference between cognate and noncognate codon-anticodon interactions.Studies of the interactions between tRNA molecules and their cognate codons in the absence of the ribosome have suggested that the triplet-triplet interaction between codon and anticodon is far too weak to account for the specificity of the tRNA selection mechanism during protein synthesis. In contrast, we suggest that such affinity measurements do not adequately describe the interaction between a codon and its cognate tRNA. Thus, such experiments can not detect conformational changes in the tRNA, and, in particular, those stabilized by the ribosome.One remarkable aspect of protein biosynthesis is its precision. Indeed, estimates of the error frequency of translation suggest that a wrong amino acid is inserted into a protein only once for every three thousand incorporated amino acids (1). Such precision has been inexplicable in terms of what is known about the mechanism of aminoacyl-tRNA selection. Thus, measurements of the affinity of trinucleotides for their complementary triplet sequences in the anti-codon of tRNA molecules indicate that such interactions are quite weak and can be characterized by equilibrium constants of the order of 103 M-1 at 00 (2-4). Furthermore, a given trinucleotide will interact with its cognate tRNA molecule with an affinity constant that is only 10 times greater than that for a related, noncognate triplet and the same tRNA. Such observations suggest that a simple triplet-triplet interaction at equilibrium can not by itself account for the fidelity of tRNA selection.Several authors have recently attempted to circumvent this problem with models that describe the tRNA selection mechanism as a nonequilibrium process (5-7). Here, the kinetics of the tRNA interaction with the codon are used to discriminate tRNA molecules through a process that must be driven by an exogenous energy supply. Nevertheless, strong, preferential binding of a specific tRNA by a mRNA-programmed ribosome in the absence of an energy source can not be explained by such mechanisms (8-10). Here, we describe a model for tRNA selection which will operate at equilibrium, and which has two additional virtues. One is that it is consistent with recent experiments concerning the interaction between tRNA and the ribosome. The other is that this model will account for a previously inexplicable class of tRNA supressor mutants.Our model is based on the following observations: In order
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