Improving the Yield and Quality of Daptomycin in Streptomyces roseosporus by Multilevel Metabolic Engineering

Abstract

Daptomycin is a cyclic lipopeptide antibiotic with a significant antibacterial action against antibiotic-resistant Gram-positive bacteria. Despite numerous attempts to enhance daptomycin yield throughout the years, the production remains unsatisfactory. This study reports the application of multilevel metabolic engineering strategies in Streptomyces roseosporus to reconstruct high-quality daptomycin overproducing strain L2797-VHb, including precursor engineering (i.e., refactoring kynurenine pathway), regulatory pathway reconstruction (i.e., knocking out negative regulatory genes arpA and phaR), byproduct engineering (i.e., removing pigment), multicopy biosynthetic gene cluster (BGC), and fermentation process engineering (i.e., enhancing O2 supply). The daptomycin titer of L2797-VHb arrived at 113 mg/l with 565% higher comparing the starting strain L2790 (17 mg/l) in shake flasks and was further increased to 786 mg/l in 15 L fermenter. This multilevel metabolic engineering method not only effectively increases daptomycin production, but can also be applied to enhance antibiotic production in other industrial strains.

Degradation mechanism of AtrA mediated by ClpXP and its application in daptomycin production in Streptomyces roseosporus

“…And due to the promotion of transcription of the BGC, the AtrA (AAAm) strain started to produce daptomycin as early as 72 h, whether in the shake flask or the bioreactor fermentation, which correspondingly prolonged the fermentation cycle. According to our previous work (Lyu et al, 2022), enhancing the transcription of dptE had little effect on the start point of daptomycin production. Thus, AtrA may also influence production of daptomycin precursors in S. roseosporus and lead to an earlier start of daptomycin production.…”

Stepwise increase of fidaxomicin in an engineered heterologous host Streptomyces albus through multi-level metabolic engineering

“…Here, VHb functions as an oxygen carrier, facilitating the proliferation of microorganisms in environments characterized by diminished levels of dissolved oxygen. The expression of VHb in a heterologous system has been demonstrated to augment the efficiency of oxygen utilization, thereby elevating the production of metabolites [ [21] , [22] , [23] ]. These series of genetic modifications were subsequently confirmed through PCR, verifying both the deletion and insertion of genes ( Fig.…”

Development of a native-locus dual reporter system for the efficient screening of the hyper-production of natural products in Streptomyces

“…Based on the genome mining of transcription factors and deciphering of regulatory mechanisms, the removal of the negative regulators ( Mao et al, 2015 ; Mao et al, 2017 ; Luo et al, 2018 ) and stabilization of the positive pleiotropic regulator AtrA ( Xu et al, 2023 ) have significantly increased daptomycin production. Meanwhile, targeting multi-level rational metabolic engineering by precursor supply, regulatory pathway reconstruction, byproduct removal, and gene cluster multiplicity have improved the yield by about 6.6 folds ( Lyu et al, 2022 ). To more comprehensively disturb the regulatory networks for the efficient increase of daptomycin production, the present study developed a dual-reporter system at the native daptomycin gene cluster and efficiently obtained a strain with improved productivity of 6.4 folds with simple UV-induced mutagenesis.…”