MassIVE MSV000093884

Partial Public

GNPS - Metabolic engineering of Streptomyces peucetius for biosynthesis of N,N-dimethylated anthracyclines

Description

Daunorubicin and doxorubicin, two anthracycline polyketides produced by Streptomyces peucetius, are potent anticancer agents that are widely used in chemotherapy, despite severe side effects. Recent advances have highlighted the potential of producing improved derivatives with reduced side effects by incorporating L-rhodosamine, the N,N-dimethyl analogue of the native amino sugar moiety. In this study, we aimed to produce N,N-dimethylated anthracyclines by engineering the doxorubicin biosynthetic pathway in the industrial S. peucetius strain GOO1. To achieve this, we introduced genes from the aclarubicin biosynthetic pathway encoding the sugar N-methyltransferases AclP and AknX2. Furthermore, the native gene for glycosyltransferase DnrS was replaced with genes encoding the aclarubicin glycosyltransferases AknS and AknT. Additionally, the gene for methylesterase RdmC from the rhodomycin biosynthetic pathway was introduced. A new host was engineered successfully, whereby genes from the aclarubicin pathway were introduced and expressed. LC-MS/MS analysis of the engineered strains showed that dimethylated sugars were efficiently produced, and that these were incorporated ino the anthracycline biosynthetic pathway to produce the novel dimethylated anthracycline N,N-dimethyldaunorubicin. Further downstream tailoring steps catalysed by the cytochrome P450 monooxygenase DoxA exhibited limited efficacy with N,N-dimethylated substrates. This resulted in only low production levels of N,N-dimethyldaunorubicin and no N,N-dimethyldoxorubicin, most likely due to the low affinity of DoxA for dimethylated substrates. S. peucetius GOO1 was engineered such as to produce N,N-dimethylated sugars, which were incorporated into the biosynthetic pathway. This allowed the successful production of N,N-dimethyldaunorubicin, an anticancer drug with reduced cytotoxicity. DoxA is the key enzyme that determines the efficiency of the biosynthesis of N,N-dimethylated anthracyclines, and engineering of this enzyme will be a major step forwards towards the efficient production of more N,N-dimethylated anthracyclines, including N,N-dimethyldoxorubicin. This study provides valuable insights into the biosynthesis of clinically relevant daunorubicin derivatives, highlighting the importance of combinatorial biosynthesis. [doi:10.25345/C5862BP1D] [dataset license: CC0 1.0 Universal (CC0 1.0)]

Keywords: doxorubicin ; anthracyclines ; anticancer ; metabolic engineering ; Streptomyces ; biosynthesis

Contact

Principal Investigators:
(in alphabetical order)
Prof. Gilles van Wezel, Leiden University, the Netherlands
Submitting User: mbhulst
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GNPS content goes here (MSV000093884 [task=19ea6d98d9964f9b831af1012401d762])
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Distinct protein accessions are counted across all files submitted in the "Statistical Analysis of Quantified Analytes" category having a "Protein" column in this dataset.

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