Biology subjects

Roberts, S. J.

Publications and source records attributed to Roberts, S. J..

4 recordsLinked to original sources

bioRxiv2023

Initial amino acid:codon assignments and strength of codon:anticodon binding

The ribosome brings 3'-aminoacyl-tRNA and 3'-peptidyl-tRNAs together to enable peptidyl transfer by binding them in two major ways. Firstly, their anticodon loops are bound to mRNA, itself anchored at the ribosomal subunit interface, by contiguous anticodon:codon pairing augmented by interactions with the decoding centre of the small ribosomal subunit. Secondly, their acceptor stems are bound by the peptidyl transferase centre which aligns the 3'-aminoacyl- and 3'-peptidyl-termini for optimal interaction of the nucleophilic amino group and electrophilic ester carbonyl group. Reasoning that intrinsic codon:anticodon binding might have been a major contributor to bringing tRNA 3'-termini into proximity at an early stage of ribosomal peptide synthesis, we wondered if primordial amino acids might have been assigned to those codons that bind the corresponding anticodon loops most tightly. By measuring the binding of anticodon stem loops to short oligonucleotides, we determined that family box codon:anticodon pairings are typically tighter than split box codon:anticodon pairings. Furthermore, we find that two family box anticodon stem loops can tightly bind a pair of contiguous codons simultaneously whereas two split box anticodon stem loops cannot. The amino acids assigned to family boxes correspond to those accessible by what has been termed cyanosulfidic chemistry, supporting the contention that these limited amino acids might have been the first used in primordial coded peptide synthesis. Table of Contents (TOC) graphic O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=64 SRC="FIGDIR/small/572171v2_ufig1.gif" ALT="Figure 1"> View larger version (13K): org.highwire.dtl.DTLVardef@189358aorg.highwire.dtl.DTLVardef@119fbeaorg.highwire.dtl.DTLVardef@e9c764org.highwire.dtl.DTLVardef@21b12c_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2023

Triplet-encoded prebiotic RNA aminoacylation

The encoding step of translation involves attachment of amino acids to cognate tRNAs by aminoacyl-tRNA synthetases, themselves the product of coded peptide synthesis. So, the question arises -- before these enzymes evolved, how were primordial tRNAs selectively aminoacylated? Here we demonstrate enzyme-free, sequence-dependent, chemoselective aminoacylation of RNA. We investigated two potentially prebiotic routes to aminoacyl-tRNA acceptor stem-overhang mimics and analyzed those oligonucleotides undergoing the most efficient aminoacylation. Overhang sequences do not influence the chemoselectivity of aminoacylation by either route. For aminoacyl-transfer from a mixed anhydride donor strand, the chemoselectivity and stereoselectivity of aminoacylation depends on the terminal three base pairs of the stem. The results support early suggestions of a second genetic code in the acceptor stem. One-Sentence SummarySelectivity of RNA stem-overhang aminoacylation is controlled by the terminal trinucleotide sequence of the stem.

molecular biology↗

bioRxiv2022

Potentially prebiotic synthesis of aminoacyl-RNA via a bridging phosphoramidate-ester intermediate

Translation according to the genetic code is made possible by selectivity both in aminoacylation of tRNA and in anticodon:codon recognition. In extant biology, tRNAs are selectively aminoacylated by enzymes using high-energy intermediates, but how this might have been achieved prior to the advent of protein synthesis has been a largely unanswered question in prebiotic chemistry. We have now elucidated a novel, prebiotically plausible stereoselective aminoacyl-RNA synthesis which starts from RNA-amino acid phosphoramidates and proceeds via phosphoramidate-ester intermediates which subsequently undergo conversion to aminoacyl-esters by mild acid hydrolysis. The chemistry avoids the intermediacy of high-energy mixed carboxy-phosphate anhydrides and is greatly favored under eutectic conditions, which also potentially allow for the requisite pH fluctuation through the variable solubility of CO2 in solid/liquid water.

biochemistry↗

bioRxiv2021

Nonenzymatic loop-closing ligation generates RNA hairpins and is a template-free way to assemble functional RNAs

RNA hairpin loops are the predominant element of secondary structure in functional RNAs. The emergence of primordial functional RNAs, such as ribozymes that fold into complex structures that contain multiple hairpin loops, is generally thought to have been supported by template-directed ligation. However, template inhibition and RNA misfolding problems impede the emergence of function. Here we demonstrate that RNA hairpin loops can be synthesized directly from short RNA duplexes with single-stranded overhangs by nonenzymatic loop-closing ligation chemistry. We show that loop-closing ligation allows full-length functional ribozymes containing a hairpin loop to be assembled free of inhibitory template strands. This approach to the assembly of structurally complex RNAs suggests a plausible pathway for the emergence of functional RNAs before a full-length RNA copying process became available.

synthetic biology↗