Methods in Enzymology
·
2019
·
IF 1.862
Wan Zhang,Linjun Zhou,Chunyu Li,Zixin Deng,Xudong Qu
Polyketides have valuable pharmaceutical properties and exhibit a high degree of structural diversity. This diversity can trace back to the polyketide synthesis assembly line, in which the gatekeeper domain ATs strictly control the selection and incorporation of the simple extender units. And thus engineering attempts targeted on ATs have been made to obtain novel polyketide skeletons, which are useful for pharmaceutical and cellular function studies. So far, method of combinatorial biosynthesis has become the most attractive and effective way, particularly through alteration of the acyltransferase (AT) specificity. Herein, upon the complex structures of a broadly selective SpnD-AT, we introduce a structure-directed protocol to manipulate the well-studied EryAT6 to broaden its substrate scope. This protocol can also be generalized to canonical AT domains engineering in the generation of novel and useful chemical probes for dissecting cellular functions.
研究领域:
生物化学
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药学
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分子生物学
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DOI
PubMed
高影响因子
Frontiers in Microbiology
·
2020
·
IF 4.236
Lü Jin,Long Qingshan,Zhao Zhilong,Chen Lu,He Weijun,Hong Jiali,Liu Kai,Wang Yemin,Pang Xiuhua,Deng Zixin,Tao Meifeng
Bacteria of the genus Saccharopolyspora produce important polyketide antibiotics, including erythromycin A (Sac. erythraea) and spinosad (Sac. spinosa). We herein report the development of an industrial erythromycin-producing strain, Sac. erythraea HOE107, into a host for the heterologous expression of polyketide biosynthetic gene clusters (BGCs) from other Saccharopolyspora species and related actinomycetes. To facilitate the integration of natural product BGCs and auxiliary genes beneficial for the production of natural products, the erythromycin polyketide synthase (ery) genes were replaced with two bacterial attB genomic integration sites associated with bacteriophages ϕC31 and ϕBT1. We also established a highly efficient conjugation protocol for the introduction of large bacterial artificial chromosome (BAC) clones into Sac. erythraea strains. Based on this optimized protocol, an arrayed BAC library was effectively transferred into Sac. erythraea. The large spinosad gene cluster from Sac. spinosa and the actinorhodin gene cluster from Streptomyces coelicolor were successfully expressed in the ery deletion mutant. Deletion of the endogenous giant polyketide synthase genes pkeA1-pkeA4, the product of which is not known, and the flaviolin gene cluster (rpp) from the bacterium increased the heterologous production of spinosad and actinorhodin. Furthermore, integration of pJTU6728 carrying additional beneficial genes dramatically improved the yield of actinorhodin in the engineered Sac. erythraea strains. Our study demonstrated that the engineered Sac. erythraea strains SLQ185, LJ161, and LJ162 are good hosts for the expression of heterologous antibiotics and should aid in expression-based genome-mining approaches for the discovery of new and cryptic antibiotics from Streptomyces and rare actinomycetes.
研究领域:
微生物学
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生物技术
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基因工程
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天然产物化学
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DOI
PubMed
高影响因子
MOLECULES
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2025
·Vol. 30
·期 17
·3474
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IF 4.6
Baoshan Li,Ke Mo,Lipeng Zhou,Yanjun Wang,Yaping Li,Wei Zhang,Chenghao Zhu,Zhirong Sun
Gastrodiae Rhizoma(tianma, TM), a traditional medicine that has food and medicine homology, faces controversy over retaining its epidermis (tianma pi, TP) during processing due to unclear phytochemical value. This study presents the first integrated approach combining GC–IMS, UHPLC–MS, andCaenorhabditis elegans(C. elegans) aging models to compare TP with the tuber core (tianma xin, TX). The results include the following: (1) A total of forty-seven volatile compounds were identified by GC–IMS, including 12 key aroma substances via relative odor activity value (ROAV ≥ 1), of which seven ((Z)-4-heptenal, β-citronellol, hexanal, 1-pentanol, 1-octen-3-one, 2-methylpropanol, and 2-butanone) were enriched in TP. (2) Non-targeted metabolomics revealed 1025 metabolites via UHPLC–MS, highlighting phenylpropanoid biosynthesis as the primary differential pathway (p< 0.05). Phenylpropanoids and polyketides exhibited predominant enrichment in TP (|log2FC| > 2, VIP > 1,p< 0.01). (3) InC. elegansmodels, TP outperformed TX in pharyngeal pumping (4.16%,p< 0.05), while both extended stress-resistant lifespan (p< 0.01). In conclusion, TP plays an essential role in establishing the characteristic odor profile of TM and retaining bioactive components, particularly phenylpropanoids. Preserving TP during processing optimally maintains the distinctive aroma profile and pharmacological value of TM, which provides valuable guidance for industrial utilization.
研究领域:
植物化学
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中医药
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代谢组学
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生药学
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衰老研究
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食药同源
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产品关联
DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）