Training old dogs to do new tricks: A general multi-pronged activation approach for natural product discovery in Actinomycetes

Tay, Dillon W. P.; Tan, Lee Ling; Heng, Elena; Zulkarnain, Nadiah; Ching, Kuan Chieh; Wibowo, Mario; Chin, Elaine J.; Tan, Zann Y.; Leong, Chung Yan; Ng, Veronica; Yang, Lay Kien; Seow, Deborah Chwee San; Koduru, Lokanand; Yoganathan, K.; Ng, Siew Bee; Lim, Yee Hwee; Wong, Fong Tian

doi:10.21203/rs.3.rs-2055033/v1

Cited by 2 publications

(4 citation statements)

References 46 publications

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“…They also demonstrated that this can be tuned by expression of fatty acyl-CoA synthase (FAS). In this work and prior observations [17], we also observed FAS to be significantly efficient in improving production of silent or low-yielding products. Additionally, similar to observations made previously with notonesomycin production in Streptomyces sp.…”

Section: Discussionsupporting

confidence: 78%

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Enhancing armeniaspirols production through multi-level engineering of a native Streptomyces producer

et al. 2023

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Background Nature has provided unique molecular scaffolds for applications including therapeutics, agriculture, and food. Due to differences in ecological environments and laboratory conditions, engineering is often necessary to uncover and utilize the chemical diversity. Although we can efficiently activate and mine these often complex 3D molecules, sufficient production of target molecules for further engineering and application remain a considerable bottleneck. An example of these bioactive scaffolds is armeniaspirols, which are potent polyketide antibiotics against gram-positive pathogens and multi-resistance gram-negative Helicobacter pylori. Here, we examine the upregulation of armeniaspirols in an alternative Streptomyces producer, Streptomyces sp. A793. Results Through an incidental observation of enhanced yields with the removal of a competing polyketide cluster, we observed seven-fold improvement in armeniaspirol production. To further investigate the improvement of armeniaspirol production, we examine upregulation of armeniaspirols through engineering of biosynthetic pathways and primary metabolism; including perturbation of genes in biosynthetic gene clusters and regulation of triacylglycerols pool. Conclusion With either overexpression of extender unit pathway or late-stage N-methylation, or the deletion of a competing polyketide cluster, we can achieve seven-fold to forty nine-fold upregulation of armeniaspirol production. The most significant upregulation was achieved by expression of heterologous fatty acyl-CoA synthase, where we observed not only a ninety seven-fold increase in production yields compared to wild type, but also an increase in the diversity of observed armeniaspirol intermediates and analogs.

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Section: Discussionsupporting

confidence: 78%

“…Additionally, similar to observations made previously with notonesomycin production in Streptomyces sp. A793 and other prior work [7,14,17], we expect that further improvement of yields can be made in regulation of nutrient sources or pleiotropic regulators.…”

Section: Discussionmentioning

confidence: 90%

Enhancing armeniaspirols production through multi-level engineering of a native Streptomyces producer

et al. 2023

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“…Cultures were then collected, freeze dried and extracted with methanol. The extracts were analysed as described in a previous study (48).…”

Section: Methodsmentioning

confidence: 99%

“…Cultures were then collected, freeze dried and extracted with methanol. The extracts were analysed as described in a previous study [49]. to Qubit ratio of 1.0 to 1.5 were used for Nanopore sequencing.…”

Section: Strainsmentioning

confidence: 99%

Cost-effective hybrid long-short read assembly delineates alternative GC-richStreptomyceschassis for natural product discovery

Heng¹,

Tan²,

Tay

et al. 2022

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With the advent of rapid automated in silico identification of biosynthetic gene clusters (BGCs), genomics presents vast opportunities to accelerate natural product (NP) discovery. However, prolific NP producers, Streptomyces, are exceptionally GC-rich (>80%) and highly repetitive within BGCs. These pose challenges in sequencing and high-quality genome assembly which are currently circumvented via intensive sequencing. Here, we outline a more cost-effective workflow using multiplex Illumina and Oxford Nanopore sequencing with hybrid long-short read assembly algorithms to generate high quality genomes. Our protocol involves subjecting long read derived assemblies to up to 4 rounds of polishing with short reads to yield accurate BGC predictions. We successfully sequenced and assembled 8 GC-rich Streptomyces genomes whose lengths range from 7.1 to 12.1 Mb at an average N50 of 5.9 Mb. Taxonomic analysis revealed previous misrepresentation among these strains and allowed us to propose a potentially new species, Streptomyces sydneybrenneri. Further comprehensive characterization of their biosynthetic, pan-genomic and antibiotic resistance features especially for molecules derived from type I polyketide synthase (PKS) BGCs reflected their potential as NP chassis. Thus, the genome assemblies and insights presented here are envisioned to serve as gateway for the scientific community to expand their avenues in NP discovery.

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Training old dogs to do new tricks: A general multi-pronged activation approach for natural product discovery in Actinomycetes

Cited by 2 publications

References 46 publications

Enhancing armeniaspirols production through multi-level engineering of a native Streptomyces producer

Enhancing armeniaspirols production through multi-level engineering of a native Streptomyces producer

Cost-effective hybrid long-short read assembly delineates alternative GC-richStreptomyceschassis for natural product discovery

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