# Chemoenzymatic Synthesis of Darobactin Antibiotics

> **NIH NIH R21** · UNIVERSITY OF CALIFORNIA LOS ANGELES · 2024 · $180,776

## Abstract

Abstract
 Darobactin A is a post-translationally modified peptide antibiotic recently isolated from photorhabdus
symbionts present in nematodes. The molecule contains a novel pattern of internal oxidative cross links that
result in its pre-organization as a beta sheet mimetic. The compound binds to the beta barrel protein BamA and
inhibits chaperone functions essential for folding of bacterial outer membrane proteins. Darobactin A is broadly
active against gram-negative pathogens in vitro and in animal models of infection.
 The dar operon encodes a single radical SAM enzyme that catalyzes formation of both oxidative cross links
observed in the natural product. The mechanism(s) behind this remarkable outcome is not yet known. We have
expressed and purified recombinant His-tagged DarE. When properly reconstituted with iron and sulfur under
anaerobic conditions, iron titration data indicates the enzyme contains three iron sulfur clusters and it rapidly
generates 5dAdo from SAM – indicating reconstituted DarE is active as a SPASM enzyme. Here we propose a
combined chemical synthesis and biosynthesis program to study DarE enzymology and to engineer semi-
synthetic forms of the natural product that can be produced on scale. Such studies hold considerable promise.
It has been nearly 60 years since a new class of antibiotics active against gram negative infections have been
developed.

## Key facts

- **NIH application ID:** 10905987
- **Project number:** 5R21AI168952-02
- **Recipient organization:** UNIVERSITY OF CALIFORNIA LOS ANGELES
- **Principal Investigator:** Patrick G. Harran
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $180,776
- **Award type:** 5
- **Project period:** 2023-08-11 → 2026-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10905987

## Citation

> US National Institutes of Health, RePORTER application 10905987, Chemoenzymatic Synthesis of Darobactin Antibiotics (5R21AI168952-02). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/10905987. Licensed CC0.

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