A diminutive and specific RNA binding site for L-tryptophan

Abstract

Selection for amino acid affinity by elution of RNAs from tryptophan–Sepharose using free L-tryptophan evokes one sequence predominantly (KD = 12 µM), a symmetrical internal loop of 3 nt per side. Though we have also isolated larger sequences with affinity for tryptophan, successively squeezed selection in randomized tracts of 70, 60, 40, 20 and 17 nt show that this internal loop is the simplest sequence that can meet the column affinity selection. From sequence variation in ∼50 independent isolates, only 26 bits of information are required to describe this loop (equivalent to only 13 fully conserved nucleotides). Thus, it is among the simplest amino acid binding sites known, as well as selective among hydrophobic side chains. Among site sequences defined as essential to affinity by conservation, protection and modification-interference, there is a recurring CCA sequence (a tryptophan anticodon triplet) which apparently forms one side of the binding site. Such conserved juxtaposition of tryptophan with a cognate coding triplet supports a stereochemical origin for the genetic code.

Synthetic RNA devices to expedite the evolution of metabolite-producing microbes

“…To demonstrate this, we chose L-tryptophan, whose naturally occurring riboswitch has not yet been discovered, as another inconspicuous target metabolite. As a starting point for the creation of a 'Tryptophan Riboselector' 16 , we selected one of the synthetic aptamers specific for L-tryptophan that are already known 33 . The readily obtained Tryptophan Riboselector (Fig.…”

Nucleotides that are essential but not conserved; a sufficient L-tryptophan site in RNA

“…This site has been independently selected 56 times in four different random-sequence backgrounds and is highly likely to be the minimal RNA structure that specifically binds the amino acid tryptophan (Majerfeld and Yarus 2005;Yarus et al 2009). We show that a combination of SELEX, molecular modeling, and testing of specific mutant constructs define an unexpected, nonconserved piece of the motif, and that incorporation of these additional constraints enabled generation of functional Trp-binding sequences in randomized contexts (Fig.…”

Imprints of the genetic code in the ribosome

“…One theory addressing this challenging question is the stereochemical hypothesis, which postulates that the genetic code developed from interactions between anticodon-or codon-containing polynucleotides and their respective amino acids (5,9). This theory is supported by RNA aptamer experiments, in which RNA molecules evolved to bind specific amino acids in vitro are enriched with anticodon and codon elements for the amino acid (6)(7)(8)10). Codons for arginine have also been found to confer binding specificity for arginine in the self-splicing group I introns (11).…”