Biology subjects

Raskar, T.

Publications and source records attributed to Raskar, T..

4 recordsLinked to original sources

bioRxiv2026

Structural adaptations for enhanced translation kinetics in evolved ribosomes

The ribosomal RNA sequence governs translation dynamics, yet understanding how changes beyond the conserved catalytic centers influence kinetics and protein yield remains limited. Using orthogonal ribosome phage-assisted continuous evolution (oRibo-PACE), we recently reported chimeric ribosomes derived from Escherichia coli, Pseudomonas aeruginosa, and Vibrio cholerae endowed with elevated orthogonal translation activity as compared to their starting counterparts. Here, we structurally characterize these kinetically enhanced ribosomes using cryo-electron microscopy and uncover a potential relationship between 16S rRNA stability and translation efficiency. Compared to their naive starting points, evolved ribosomes exhibit extensive RNA structural adaptation, often introduced by mismatches at key helical junctions, which leads to local RNA-protein rearrangements and destabilizes non-canonical base pairs. Compensatory mutations that restore base-pairing stability and eliminate flexibility reduced translational activity to wild-type levels. Across trajectories, increased translational output correlates with subtle, localized changes in the 16S rRNA sequence that introduce limited structural destabilization at specific elements. Taken together, our work provides new insights into rRNA structural malleability and establishes principles for engineering ribosomes with altered translation properties. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=74 SRC="FIGDIR/small/706023v2_ufig1.gif" ALT="Figure 1"> View larger version (16K): org.highwire.dtl.DTLVardef@8678bdorg.highwire.dtl.DTLVardef@60cde5org.highwire.dtl.DTLVardef@16db00dorg.highwire.dtl.DTLVardef@413ed5_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2026

Structural modification of oxazolidinone antibiotics alters nascent peptide stalling preference and peptide trajectory through the ribosome

The oxazolidinone antibiotic linezolid binds to the peptidyl transferase center of the ribosome, where it inhibits a subset of peptide bond formation events. This context-specificity of translation inhibition is dictated by the nature of the amino acid at the penultimate position of the nascent peptide. It remains unknown whether this is a general feature of oxazolidinones and whether it can be modulated by their structural alterations. Here, we show that the oxazolidinone tedizolid also inhibits translation in a context-specific manner, but with dramatically altered selectivity, favoring Ile, His, and Gln as the penultimate residues. Delpazolid, which shares the C5 hydroxymethyl moiety with tedizolid, shows a similar preference. Structural analysis of the ribosome with tedizolid and a stalled nascent peptide showed a compacted, helical conformation of the nascent chain induced by the drug. Our findings reveal that stalling preferences of oxazolidinones can be modulated by structural modifications within this antibiotic class.

biochemistry↗

bioRxiv2024

Context-specific inhibition of mitochondrial ribosomes by phenicol and oxazolidinone antibiotics

The antibiotics chloramphenicol (CHL) and oxazolidinones including linezolid (LZD) are known to inhibit mitochondrial translation. This can result in serious, potentially deadly, side effects when used therapeutically. Although the mechanism by which CHL and LZD inhibit bacterial ribosomes has been elucidated in detail, their mechanism of action against mitochondrial ribosomes has yet to be explored. CHL and oxazolidinones bind to the ribosomal peptidyl transfer center (PTC) of the bacterial ribosome and prevent incorporation of incoming amino acids under specific sequence contexts, causing ribosomes to stall only at certain sequences. Through mitoribosome profiling, we show that inhibition of mitochondrial ribosomes is similarly context-specific - CHL and LZD lead to mitoribosome stalling primarily when there is an alanine, serine, or threonine in the penultimate position of the nascent peptide chain. We further validate context-specific stalling through in vitro translation assays. A high resolution cryo-EM structure of LZD bound to the PTC of the human mitoribosome shows extensive similarity to the mode of bacterial inhibition and also suggests potential avenues for altering selectivity. Our findings could help inform the rational development of future, less mitotoxic, antibiotics, which are critically needed in the current era of increasing antimicrobial resistance.

biochemistry↗

bioRxiv2020

Diffusive dynamics of Aspartate α-decarboxylase (ADC) liganded with D-serine in aqueous solution

Incoherent neutron spectroscopy, in combination with dynamic light scattering was used to investigate the effect of ligand binding on the center-of-mass self-diffusion and internal diffusive dynamics of E.coli aspartate -decarboxylase (ADC). The X-ray crystal structure of the D-serine inhibitor complex with ADC was also determined, and molecular dynamics simulations used to further probe the structural rearrangements that occur as a result of ligand binding. These experiments reveal the existence of higher order oligomers of the ADC tetramer on ns-ms time-scales, and also show that ligand binding both affects the ADC internal diffusive dynamics and appears to further increase the size of the higher order oligomers.

biophysics↗