On the physical basis for ambiguity in genetic coding interactions.

Abstract

We report the relative stabilities, in the form of complex lifetimes, of complexes between two tRNAs complementary, or nearly so, in their anticodons. The results show striking parallels with the genetic coding rules, including the wobble interaction and the role of modified nucleotides sAU and V (a 5-oxyacetic acid derivative of U). One important difference between the genetic code and the pairing rules in the tRNAtRNA interaction is the stability in the latter of the short wobble pairs, which the wobble hypothesis excludes. We stress the potential of U for translational errors, and suggest a simple stereochemical basis for ribosome-mediated discrimination against short wobble pairs. Surprisingly, the stability of anticodon-anticodon complexes does not vary systematically with GC composition, unlike all other known double helices. How (10)(11)(12)(13)(14). Therefore, we selected this model system for more intensive study of base pairing rules; the observed strong parallels with the genetic coding rules justify our choice a posteriori."Recognition" between two tRNA anticodon loops can be expressed quantitatively by the association equilibrium constant, or by the kinetic constants for the association-dissociation reaction. Because the association rate constants are found to vary only moderately (by about a factor of five; unpublished results), the association equilibrium constant is approximately inversely proportional to the dissociation rate constant. Hence, we have selected a single parameter, the lifetime of the anticodonanticodon complex, determined by the temperature-jump technique, as a measure of the strength of association and therefore of the extent of recognition.In general we find that a "correct" base triplet interaction (as determined by the genetic coding rules) has a long lifetime. However, the lifetime of correct complexes frequently is not more than two orders of magnitude longer than the lifetime of some of the incorrect complexes that have mismatching bases. Therefore, to avoid errors, the ribosome must amplify the small difference in lifetime to produce a greater distinction between correct and incorrect pairings in protein synthesis. Specific mechanisms for such an amplification have been discussed under the heading of kinetic proofreading (15)(16)(17). MATERIALS AND METHODSThe procedures used in this work were described in a preceding paper (14). The concentration of each tRNA was 1.3-1.8 ,uM, based on amino acid acceptance activity. Temperature jumps of up to 100 were used, and relaxations were monitored at 266 nm. Control experiments on single samples of tRNA (not mixed with their complement) showed no slow relaxation signals between 00 and 150, except as reported explicitly below.The lifetimes (T*) of anticodon-anticodon complexes at 90 were determined from the relaxation time r, using the equation

Queuosine modification of the wobble base in tRNAHis influences ‘in vivo’ decoding properties.

“…This is in agreement with the fact (Noguchi et al, 1982) that the presence of the Q-base enhances the efficiency of tRNATYr binding to both tyrosine codons. Our results, indicating an influence of the Q-base on the decoding process, are further supported by ribosome triplet binding experiments of Hatfield et al (1982) and by observations concerning the stability of complexes formed between two tRNA anticodons (Grosjean et al, 1978). From the results of the latter approach it can be concluded that the hypermodified base Q reduces the difference in the binding energy to the two complementary sequences NAU and NAC.…”

Possible conformations of double-helical polynucleotides containing incorrect base pairs

“…Therefore our results cannot be compared directly with the experimental data on interaction of two tRNAs having mutually complementary or almost complementary anticodons (25). The authors of paper (25) suggest a considerable change of dihedral angles of the sugar-phosphate backbone upon formation of pyrimidine-pyrimidine pairs.…”

A code in the protein coding genes

“…A stacked configuration pairs with a five base codon. Note: a primitive codon -anticodon pairing involving only three bases such as an actual one without ribosome is not stable enough for codon translation (Grosjean et al, 1978). The flip mechanism (detailed below for the RNY model) between these two stacked configurations leads to primitive protein genes consisting of a series of RRY codons (RRY model) (Crick et al, 1976).…”