Is There a Discriminator Site in Transfer RNA?

Abstract

We determined the nature of the fourth nucleotide from the 3'-end of several Escherichia coli tRNAs, and tabulated these results with the same data for all known tRNA sequences. We find a striking constancy of the fourth nucleotide in tRNAs specific for a given amino acid. Furthermore, tRNAs specific for chemically related amino acids are very likely to have the same nucleotide at the fourth position.One possible explanation for these regularities is the "discriminator" hypothesis: The code by which tRNA is recognized by its cognate aminoacyl-tRNA synthetase is logically hierarchical, with the fourth nucleotide serving as a primary "discriminator" site to subdivide the tRNAs into groups for recognition purposes. Each such group could have its own recognition code, or could be further subdivided by a secondary discriminator site. According to this hypothesis, chemically similar amino acids have the same discriminator nucleotide because they evolved from a single set of related amino acids indistinguishable to a primitive system.There are other possible explanations for the observed regularities at the fourth nucleotide. For example, it is conceivable that the position is used for a direct physical interaction with the amino acid in the charging process, and chemically similar amino acids naturally select the same nucleotide. Further experiments can be expected to clarify this question.The specific recognition of a protein by a nucleic-acid macromolecule is a well studied, but still poorly understood, process. One of the best model systems for this process is the proper recognition of tRNA by the cognate aminoacyl-tRNA synthetase. This recognition must be remarkably specific in order for the enzyme to discriminate between structurally very similar tRNA molecules so as to ensure the correct translation of the genetic message. Although the great advances in the methodology of sequencing nucleic acids have provided much detailed insight into tRNA primary structure, no clear ideas on which features of the tRNA molecule are recognized by the enzyme have emerged. Indeed, a multitude of observations that are seemingly contradictory from species to species raise doubts whether there is a uniform recognition mechanism for all tRNAs (1). All this, of course, is related to the tertiary structure of tRNA which, at present, is not understood.Since the tRNAs specialize in recognizing with a high degree of fidelity very many different codons, one of the early thoughts on recognition attributed specificity to the anticodon region (2). This general notion was specifically contradicted by evidence on suppressor tRNA (3) and the fact that tRNAs with seemingly unrelated codons (e.g., serine) are recognized by one enzyme (4). Studies onmodification of tRNA by UV irradiation led to the proposal that the three last base-pairs in the amino-acid acceptor stem are the specific recognition site (5). That this simple proposal was also not universally applicable was soon evident; for instance, from studies with isoacceptor tRNAs from ...

Specific valylation identity of turnip yellow mosaic virus RNA by yeast valyl‐tRNA synthetase is directed by the anticodon in a kinetic rather than affinity‐based discrimination

“…The position adjacent to the 3'-CCA end in tRNAs has long been proposed to participate in the discrimination of tRNA by aminoacyl-tRNA synthetases, and has come to be known as the 'discriminator base' [47]. Recent results on the aminoacylation of tRNA variants with substitutions at this position have supplied some confirmation to this view [48,49].…”

tRNA acceptor-stem and anticodon bases embed separate features of amino acid chemistry

“…25 Here, 0 represents ambiguity associated with the first base, which can be either G or A. Acceptor stem coding proceeds in analogous fashion for the 8 bases in the stem plus the unpaired "discriminator" base D (usually base number 73 3 ). 26 The two aaRS Classes generally recognize opposite faces of tRNA interacting with either the major or minor groove. 27 Omission of the distinction made possible by these different modes of recognition seriously degraded the performance of both acceptor stem and anticodon models.…”

U2 Small Nuclear RNA Is a Substrate for the CCA-adding Enzyme (tRNA Nucleotidyltransferase)

“…In fact, the 3Ј end of U2 strongly resembles the minimal substrate for the CCA-adding enzyme: a tRNA minihelix (the "top half "of tRNA) in which the acceptor stem stacks on the TC stem/loop. Not only is U2 stem IV 12 bp, the optimal length for CCA addition by the E. coli and S. shibatae enzymes to a minihelix substrate (12), but the stem is followed by A 185 CCA OH , where A 185 is formally analogous to the discriminator base of tRNA, most frequently a purine (74). Thus, in principle, the CCA-adding enzyme might be capable of completely rebuilding the 3Ј-terminal CCA sequence of U2 snRNA if nucleases overprocessed the U2 snRNA precursor or other 3Ј exonucleases attacked the mature U2 snRNA.…”