Establishing a New Methodology for Genome Mining and Biosynthesis of Polyketides and Peptides through Yeast Molecular Genetics

• DOI: 10.1002/cbic.201100798
• Publication Date: 2012-03-24T08:48:44Z
• Authors: Kan'ichiro Ishiuchi; Takehito Nakazawa; Takashi Ookuma; Satoru Sugimoto; Michio Sato; Yuta Tsunematsu; Noriyasu Ishikawa; Hiroshi Noguchi; Kinya Hotta; Hisao Moriya; Kenji Watanabe
• Keywords: Nonribosomal peptide; Polyketide synthase; Heterologous expression

Abstract Fungal genome sequencing has revealed many genes coding for biosynthetic enzymes, including polyketide synthases and nonribosomal peptide synthetases. However, characterizing these enzymes identifying the compounds they synthesize remains a challenge, whether are expressed in their original hosts or more tractable heterologous hosts, such as yeast. Here, we developed streamlined method isolating from fungal sources producing bioactive molecules an engineered Saccharomyces cerevisiae host strain. We used overlap extension PCR yeast homologous recombination to clone desired synthase synthetase (5–20 kb) into expression vector quickly efficiently. This approach was successfully five one synthetase, various species. Subsequent detailed chemical characterizations of resulting natural products identified six two products, which new compound. Our system should facilitate investigating uncharacterized genes, novel rationally engineering pathways production enzyme analogues possessing modified bioactivity.