lycodiversification of gentamicins through in vivo glycosyltransferase swapping enabled the creation of novel hybrid aminoglycoside antibiotics with potent activity and low ototoxicity


Xinyun Jian#, Cheng Wang#, Shijuan Wu#, Guo Sun, Chuan Huang, Chengbing Qiu, Yuanzheng Liu, Peter F. Leadlay, Dong Liu, Zixin Deng, Fuling Zhou, and Yuhui Sun*


Acta Pharmaceutica Sinica B 2024

Epub Date: 3 May 2024

DOI: 10.1016/j.apsb.2024.04.030

Abstract

Aminoglycosides (AGs) are a class of antibiotics with a broad spectrum of activity. However, their use is limited by safety concerns associated with nephrotoxicity and ototoxicity, as well as drug resistance. To address these issues, semi-synthetic approaches for modifying natural AGs have generated new generations of AGs, however, with limited types of modification due to significant challenges in synthesis. This study explores a novel approach that harness the bacterial biosynthetic machinery of gentamicins and kanamycins to create hybrid AGs. This was achieved by glycodiversification of gentamicins via swapping the glycosyltransferase (GT) in their producer with the GT from kanamycins biosynthetic pathway and resulted in the creation of a series of novel AGs, therefore referred to as genkamicins (GKs). The manipulation of the hybrid metabolic pathway enabled the target accumulation of different GK species and the successful isolation and characterization of six GK components. These compounds display retained antimicrobial activity against a panel of World Health Organization (WHO) critical priority pathogens, and GK-C2a, in particular, demonstrates low ototoxicity compared to clinical drugs in zebrafish embryos. This study provides a new strategy for diversifying the structure of AGs and a potential avenue for developing less toxic AG drugs to combat infectious diseases.

Engineered cytosine base editor enabling broad-scope and high-fidelity gene editing in Streptomyces


Jian Wang, Ke Wang, Zhe Deng, Zhiyu Zhong, Guo Sun, Qing Mei, Fuling Zhou, Zixin Deng, and Yuhui Sun*


Nature Communications 2024, 15:5687

Epub Date: 7 July 2024

DOI: 10.1038/s41467-024-49987-3




Abstract

Base editing (BE) faces protospacer adjacent motif (PAM) constraints and off-target effects in both eukaryotes and prokaryotes. For Streptomyces, renowned as one of the most prolific bacterial producers of antibiotics, the challenges are more pronounced due to its diverse genomic content and high GC content. Here, we develop a base editor named eSCBE3-NG-Hypa, tailored with both high-efficiency and -fidelity for Streptomyces. Of note, eSCBE3-NG-Hypa recognizes NG PAM and exhibits high activity at challenging sites with high GC content or GC motifs, while displaying minimal off-target effects. To illustrate its practicability, we employ eSCBE3-NG-Hypa to achieve precise key amino acid conversion of the dehydratase (DH) domains within the modular polyketide synthase (PKS) responsible for the insecticide avermectins biosynthesis, achieving domains inactivation. The resulting DH-inactivated mutants, while ceasing avermectins production, produce a high yield of oligomycin, indicating competitive relationships among multiple biosynthetic gene clusters (BGCs) in Streptomyces avermitilis. Leveraging this insight, we use eSCBE3-NG-Hypa to introduce premature stop codons into competitor gene cluster of ave in an industrial S. avermitilis, with the mutant Δolm exhibiting the highest 4.45-fold increase in avermectin B1a compared to the control. This work provides a potent tool for modifying biosynthetic pathways and advancing metabolic engineering in Streptomyces.