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A BioBricks Metabolic Engineering Platform for the Biosynthesis of Anthracyclinones in Streptomyces coelicolor

Abstract

Actinomycetes produce a variety of clinically indispensable molecules, such as antineoplastic anthracyclines. However, the actinomycetes are hindered in their further development as genetically engineered hosts for the synthesis of new anthracycline analogues due to their slow growth kinetics associated with their mycelial life cycle and the lack of a comprehensive genetic toolbox for combinatorial biosynthesis. In this report, we tackled both issues via the development of the BIOPOLYMER (BIOBricks POLYketide Metabolic EngineeRing) toolbox: a comprehensive synthetic biology toolbox consisting of engineered strains, promoters, vectors, and biosynthetic genes for the synthesis of anthracyclinones. An improved derivative of the production host Streptomyces coelicolor M1152 was created by deleting the matAB gene cluster that specifies extracellular poly-β-1,6-N-acetylglucosamine (PNAG). This resulted in a loss of mycelial aggregation, with improved biomass accumulation and anthracyclinone production. We then leveraged BIOPOLYMER to engineer four distinct anthracyclinone pathways, identifying optimal combinations of promoters, genes, and vectors to produce aklavinone, 9-epi-aklavinone, auramycinone, and nogalamycinone at titers between 15–20 mg/L. Optimization of nogalamycinone production strains resulted in titers of 103 mg/L. We structurally characterized six anthracyclinone products from fermentations, including new compounds 9,10-seco-7-deoxy-nogalamycinone and 4-O-β-d-glucosyl-nogalamycinone. Lastly, we tested the antiproliferative activity of the anthracyclinones in a mammalian cancer cell viability assay, in which nogalamycinone, auramycinone, and aklavinone exhibited moderate cytotoxicity against several cancer cell lines. We envision that BIOPOLYMER will serve as a foundational platform technology for the synthesis of designer anthracycline analogues.
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Category

Academic article

Language

English

Author(s)

  • Rongbin Wang
  • Jennifer Nguyen
  • Jacob Hecht
  • Nora Schwartz
  • Katelyn V. Brown
  • Larissa V. Ponomareva
  • Magdalena Niemczura
  • Dino van Dissel
  • Gilles P. Van Wezel
  • Jon S. Thorson
  • Mikko Metsä-Ketelä
  • Khaled A. Shaaban
  • S. Eric Nybo

Affiliation

  • SINTEF Industry / Biotechnology and Nanomedicine
  • University of Turku
  • Leiden University
  • University of Kentucky
  • Ferris State University

Year

2022

Published in

ACS Synthetic Biology

Volume

11

Issue

12

Page(s)

4193 - 4209

View this publication at Norwegian Research Information Repository