# Elucidating the role of context-specificity in oxazolidinone mechanism of action

> **NIH NIH F31** · UNIVERSITY OF CALIFORNIA, SAN FRANCISCO · 2024 · $45,909

## Abstract

PROJECT SUMMARY/ABSTRACT
Antimicrobial resistance is an urgent and growing threat, with a projected toll of 10 million deaths annually by
2050. An improved understanding of the mechanisms of existing antibiotics, applied in the development of
improved drugs capable of circumventing common resistance mechanisms, will be key towards combating this
ongoing crisis. A class of fully synthetic ribosome-binding antibiotics, oxazolidinones are used clinically as a
last defense against multidrug resistant gram-positive pathogens. Though these molecules have historically
been considered global translation inhibitors, recent applications of ribosome profiling and cryo-EM have
shown that clinical frontrunner linezolid in fact exhibits sequence-specific inhibition of translation. Although
other classes of antibiotics have been shown to exhibit context-specific stalling, it remains unclear how and to
what extent context-specificity contributes to antibiotic mechanism of action. The synthetic origin of
oxazolidinone antibiotics has privileged them with status as a last-resort treatment for especially difficult
infections, due to the decreased frequency of resistance compared to their natural product counterparts. As
such, the recent discovery of multiple oxazolidinone resistance factors has been especially alarming. One of
these proteins, Cfr, confers resistance to eight classes of antibiotic, including linezolid, through methylation of a
single rRNA residue. The clinical oxazolidinones exhibit variable activity against this modification, however;
while linezolid is highly susceptible to Cfr-mediated resistance, tedizolid maintains activity in Cfr-expressing
strains. This proposal will explore 1) the mechanisms by which tedizolid avoids Cfr-mediated resistance and 2)
the interconnectivity of context-specific stalling and overall antibiotic efficacy in the oxazolidinone class of
antibiotics. In the first aim, I will use ribosome profiling and cryo-EM in concert with MIC assays for efficacy to
explore the difference in context-selectivity between tedizolid and the parent compound, as well as in the Cfr-
modified context. My 2nd aim will utilize an in vitro ribosome profiling assay in addition to cryo-EM to perform a
detailed analysis of the structure-activity relationship between oxazolidinone context-specific stalling and
antibiotic efficacy. Over the course of these two aims, I will investigate the model that context-specific stalling is
integral to oxazolidinone function as an antibiotic and evaluate the effects of structural modification on said
activity. I hypothesize that key structural differences induce a difference in context-specific stalling between the
two clinical oxazolidinones which additionally contributes to tedizolid avoidance of Cfr-mediated resistance.
This proposal is highly relevant to NIAID’s missions in understanding and treatment of infectious disease. The
research will provide significant insight into the mechanism of action of a key cla...

## Key facts

- **NIH application ID:** 10827332
- **Project number:** 1F31AI181568-01
- **Recipient organization:** UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
- **Principal Investigator:** Jordan Isabelle Kleinman
- **Activity code:** F31 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $45,909
- **Award type:** 1
- **Project period:** 2024-09-01 → 2026-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10827332

## Citation

> US National Institutes of Health, RePORTER application 10827332, Elucidating the role of context-specificity in oxazolidinone mechanism of action (1F31AI181568-01). Retrieved via AI Analytics 2026-05-26 from https://api.ai-analytics.org/grant/nih/10827332. Licensed CC0.

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