Search NASA⌕ Search

Engineering topics

Nguyen, Ha An

Publications and source records attributed to Nguyen, Ha An.

3 records
All sourcesNASA NTRSDOE OSTI
DOE OSTI2023

Antibiotic that inhibits trans -translation blocks binding of EF-Tu to tmRNA but not to tRNA

trans-Translation is conserved throughout bacteria and is essential in many species. High-throughput screening identified a tetrazole-based trans-translation inhibitor, KKL-55, that has broad-spectrum antibiotic activity. A biotinylated version of KKL-55 pulled down elongation factor thermo-unstable (EF-Tu) from bacterial lysates. Purified EF-Tu bound KKL-55 in vitro with a K d = 2 µM, confirming a high-affinity interaction. An X-ray crystal structure showed that KKL-55 binds in domain 3 of EF-Tu, and mutation of residues in the binding pocket abolished KKL-55 binding. RNA-binding assays in vitro showed that KKL-55 inhibits binding between EF-Tu and transfer-messenger RNA (tmRNA) but not between EF-Tu and tRNA. These data demonstrate a new mechanism for the inhibition of EF-Tu function and suggest that this specific inhibition of EF-Tu•tmRNA binding is a viable target for antibiotic development.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Disruption of evolutionarily correlated tRNA elements impairs accurate decoding

Bacterial transfer RNAs (tRNAs) contain evolutionarily conserved sequences and modifications that ensure uniform binding to the ribosome and optimal translational accuracy despite differences in their aminoacyl attachments and anticodon nucleotide sequences. In the tRNA anticodon stem-loop, the anticodon sequence is correlated with a base pair in the anticodon loop (nucleotides 32 and 38) to tune the binding of each tRNA to the decoding center in the ribosome. Disruption of this correlation renders the ribosome unable to distinguish correct from incorrect tRNAs. The molecular basis for how these two tRNA features combine to ensure accurate decoding is unclear. In this work, we solved structures of the bacterial ribosome containing either wild-type tRNA$^{\mathrm{Ala}}_{\mathrm{GGC}}$ or tRNA$^{\mathrm{Ala}}_{\mathrm{GGC}}$ containing a reversed 32–38 pair on cognate and near-cognate codons. Structures of wild-type tRNA$^{\mathrm{Ala}}_{\mathrm{GGC}}$ bound to the ribosome reveal 23S ribosomal RNA (rRNA) nucleotide A1913 positional changes that are dependent on whether the codon-anticodon interaction is cognate or near cognate. Further, the 32–38 pair is destabilized in the context of a near-cognate codon-anticodon pair. Reversal of the pairing in tRNA$^{\mathrm{Ala}}_{\mathrm{GGC}}$ ablates A1913 movement regardless of whether the interaction is cognate or near cognate. These results demonstrate that disrupting 32–38 and anticodon sequences alters interactions with the ribosome that directly contribute to misreading.

59 BASIC BIOLOGICAL SCIENCES↗