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

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 biosynthetic pathway enabled the targeted accumulation of different GK species and the 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.

Analyzing and engineering of the biosynthetic pathway of mollemycin A for enhancing its production

Mollemycin A(MOMA)is a unique glyco-hexadepsipeptide-polyketide that was isolated from a Streptomyces sp.derived from the Australian marine environment.MOMA exhibits remarkable inhibitory activity against both drug-sensitive and multidrug-resistant malaria parasites.Optimizing MOMA through structural modifications or product enhancements is necessary for the development of effective analogues.However,modifying MOMA using chemical approaches is challenging,and the production titer of MOMA in the wild-type strain is low.This study identified and characterized the biosynthetic gene cluster of MOMA for the first time,proposed its complex biosynthetic pathway,and achieved an effective two-pronged enhancement of MOMA production.The fermentation medium was optimized to increase the yield of MOMA from 0.9 mg L^(-1)to 1.3 mg L^(-1),a 44%boost.Additionally,a synergistic mutant strain was developed by deleting the momB3 gene and overexpressing momB2,resulting in a 2.6-fold increase from 1.3 mg L^(-1)to 3.4 mg L^(-1).These findings pave the way for investigating the biosynthetic mechanism of MOMA,creating opportunities to produce a wide range of MOMA analogues,and developing an efficient strain for the sustainable and economical production of MOMA and its analogues.

Advances in engineering methylotrophic yeast for biosynthesis of valuable chemicals from methanol

Methylotrophic yeasts and bacteria, which can use methanol as carbon and energy source, have beenwildly used as microbial cell factories for biomanufacturing. Due to their robustness in industrial harshconditions, methylotrophic yeasts such as Pichia pastoris have been explored as a cell factory forproduction of proteins and high-value chemicals. Methanol utilization pathway （MUT） is highlyregulated for efficient methanol utilization, and the downstream pathways need extensively constructedand optimized toward target metabolite biosynthesis. Here, we present an overview of methanolmetabolism and regulation in methylotrophic yeasts, among which we focus on the regulation of keygenes involved in methanol metabolism. Besides, the recent progresses in construction and optimizationof downstream biosynthetic pathways for production of high value chemicals, such as polyketides, fattyacids and isoprenoids, are further summarized. Finally, we discuss the current challenges and feasiblestrategies toward constructing efficient methylotrophic cell factories may promote wide applications inthe future.