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Interstrand Aminoacyl Transfer in a tRNA Acceptor Stem-Overhang Mimic.

ABSTRACT: Protein-catalyzed aminoacylation of the 3'-overhang of tRNA by an aminoacyl-adenylate could not have taken place prior to the advent of genetically coded peptide synthesis, and yet the latter process has an absolute requirement for aminoacyl-tRNA. There must therefore have been an earlier nonprotein-catalyzed means of generating aminoacyl-tRNA. Here, we demonstrate efficient interstrand aminoacyl transfer from an aminoacyl phosphate mixed anhydride at the 5'-terminus of a tRNA acceptor stem mimic to the 2',3'-diol terminus of a short 3'-overhang. With certain five-base 3'-overhangs, the transfer of an alanyl residue is highly stereoselective with the l-enantiomer being favored to the extent of ∼10:1 over the d-enantiomer and is much more efficient than the transfer of a glycyl residue. N-Acyl-aminoacyl residues are similarly transferred from a mixed anhydride with the 5'-phosphate to the 2',3'-diol but with a different dependence of efficiency and stereoselectivity on the 3'-overhang length and sequence. Given a prebiotically plausible and compatible synthesis of aminoacyl phosphate mixed anhydrides, these results suggest that RNA molecules with acceptor stem termini resembling modern tRNAs could have been spontaneously aminoacylated, in a stereoselective and chemoselective manner, at their 2',3'-diol termini prior to the onset of protein-catalyzed aminoacylation.

SUBMITTER: Wu LF

PROVIDER: S-EPMC8397310 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Summary Aminoacyl-tRNA synthetases (aaRSs) serve a dual role in charging tRNAs. Their enzymatic activities both provide protein synthesis flux and reduce uncharged tRNA levels. Although uncharged tRNAs can negatively impact bacterial growth, substantial concentrations of tRNAs remain deacylated even under nutrient-rich conditions. Here we show that tRNA charging in Bacillus subtilis is not maximized due to optimization of aaRS production during rapid growth, which prioritizes demands in protein synthesis over charging levels. The presence of uncharged tRNAs is alleviated by precisely tuned translation kinetics and the stringent response, both insensitive to aaRS overproduction but sharply responsive to underproduction, allowing for just-enough aaRS production atop a “fitness cliff”. Notably, we find that the stringent response mitigates fitness defects at all aaRS underproduction levels even without external starvation. Thus, adherence to minimal, flux-satisfying protein production drives limited tRNA charging and provides a basis for the sensitivity and setpoints of an integrated growth-control network.

2020-07-20 | GSE141389 | GEO

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