title: Genetic engineering and heterologous expression of the disorazol biosynthetic gene cluster via Red/ET recombineering.
abstract: Disorazol, a macrocyclic polykitide produced by the myxobacterium Sorangium cellulosum So ce12 and it is reported to have potential cytotoxic activity towards several cancer cell lines, including multi-drug resistant cells. The disorazol biosynthetic gene cluster (dis) from Sorangium cellulosum (So ce12) was identified by transposon mutagenesis and cloned in a bacterial artificial chromosome (BAC) library. The 58-kb dis core gene cluster was reconstituted from BACs via Red/ET recombineering and expressed in Myxococcus xanthus DK1622. For the first time ever, a myxobacterial trans-AT polyketide synthase has been expressed heterologously in this study. Expression in M. xanthus allowed us to optimize the yield of several biosynthetic products using promoter engineering. The insertion of an artificial synthetic promoter upstream of the disD gene encoding a discrete acyl transferase (AT), together with an oxidoreductase (Or), resulted in 7-fold increase in disorazol production. The successful reconstitution and expression of the genetic sequences encoding for these promising cytotoxic compounds will allow combinatorial biosynthesis to generate novel disorazol derivatives for further bioactivity evaluation.
authors.affiliation: Department of Microbial Natural Products, Helmholtz Institute for Pharmaceutical Research Saarland, Helmholtz Centre for Infection Research and Department of Pharmaceutical Biotechnology, Saarland University, Campus E8.1, 66123 Saarbrücken, Germany.|Shandong University - Helmholtz Joint Institute of Biotechnology, State Key Laboratory of Microbial Technology, School of Life Science, Shandong University, Jinan 250100, People's Republic of China.
authors.forename: Qiang
authors.initials: Q
authors.lastname: Tu
authors.affiliation: Department of Microbial Natural Products, Helmholtz Institute for Pharmaceutical Research Saarland, Helmholtz Centre for Infection Research and Department of Pharmaceutical Biotechnology, Saarland University, Campus E8.1, 66123 Saarbrücken, Germany.
authors.forename: Jennifer
authors.initials: J
authors.lastname: Herrmann
authors.affiliation: College of Life Science, Key Laboratory of Microbial Molecular Biology of Hunan Province, Hunan Normal University, Changsha 410081, Hunan Province, People's Republic of China.
authors.forename: Shengbiao
authors.initials: S
authors.lastname: Hu
authors.affiliation: Department of Microbial Natural Products, Helmholtz Institute for Pharmaceutical Research Saarland, Helmholtz Centre for Infection Research and Department of Pharmaceutical Biotechnology, Saarland University, Campus E8.1, 66123 Saarbrücken, Germany.
authors.forename: Ritesh
authors.initials: R
authors.lastname: Raju
authors.affiliation: Shandong University - Helmholtz Joint Institute of Biotechnology, State Key Laboratory of Microbial Technology, School of Life Science, Shandong University, Jinan 250100, People's Republic of China.
authors.forename: Xiaoying
authors.initials: X
authors.lastname: Bian
authors.affiliation: Shandong University - Helmholtz Joint Institute of Biotechnology, State Key Laboratory of Microbial Technology, School of Life Science, Shandong University, Jinan 250100, People's Republic of China.
authors.forename: Youming
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Department of Microbial Natural Products, Helmholtz Institute for Pharmaceutical Research Saarland, Helmholtz Centre for Infection Research and Department of Pharmaceutical Biotechnology, Saarland University, Campus E8.1, 66123 Saarbrücken, Germany.
authors.forename: Rolf
authors.initials: R
authors.lastname: Müller
mesh_terms: D053898:Biosynthetic Pathways / Q000235:genetics*; D045744:Cell Line, Tumor / Q000187:drug effects; D022202:Chromosomes, Artificial, Bacterial / Q000187:drug effects; D004926:Escherichia coli / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D005818:Genetic Engineering; D006801:Humans; D005810:Multigene Family; D016296:Mutagenesis; D016941:Myxococcus xanthus / Q000235:genetics / Q000378:metabolism; D010080:Oxazoles / Q000378:metabolism; D011995:Recombination, Genetic
chemical_list: D010080:Oxazoles; C092171:disorazol A