title: Heterologous production of kasugamycin, an aminoglycoside antibiotic from Streptomyces kasugaensis, in Streptomyces lividans and Rhodococcus erythropolis L-88 by constitutive expression of the biosynthetic gene cluster.
abstract: Kasugamycin (KSM), an aminoglycoside antibiotic isolated from Streptomyces kasugaensis cultures, has been used against rice blast disease for more than 50 years. We cloned the KSM biosynthetic gene (KBG) cluster from S. kasugaensis MB273-C4 and constructed three KBG cassettes (i.e., cassettes I-III) to enable heterologous production of KSM in many actinomycetes by constitutive expression of KBGs. Cassette I comprised all putative transcriptional units in the cluster, but it was placed under the control of the Pneopromoter from Tn5. It was not maintained stably in Streptomyces lividans and did not transform Rhodococcus erythropolis. Cassette II retained the original arrangement of KBGs, except that the promoter of kasT, the specific activator gene for KBG, was replaced with PrpsJ, the constitutive promoter of rpsJ from Streptomyces avermitilis. To enhance the intracellular concentration of myo-inositol, an expression cassette of ino1 encoding the inositol-1-phosphate synthase from S. avermitilis was inserted into cassette II to generate cassette III. These two cassettes showed stable maintenance in S. lividans and R. erythropolis to produce KSM. Particularly, the transformants of S. lividans induced KSM production up to the same levels as those produced by S. kasugaensis. Furthermore, cassette III induced more KSM accumulation than cassette II in R. erythropolis, suggesting an exogenous supply of myo-inositol by the ino1 expression in the host. Cassettes II and III appear to be useful for heterologous KSM production in actinomycetes. Rhodococcus exhibiting a spherical form in liquid cultivation is also a promising heterologous host for antibiotic fermentation.
authors.affiliation: Department of Biotechnology, Akita Prefectural University, 241-438 Kaidobata-Nishi, Akita City, Nakano Shimoshinjo, 010-0195, Japan. ccasga@akita-pu.ac.jp.
authors.forename: Kano
authors.identifier: http://orcid.org/0000-0002-2781-8190
authors.initials: K
authors.lastname: Kasuga
authors.affiliation: Department of Biotechnology, Akita Prefectural University, 241-438 Kaidobata-Nishi, Akita City, Nakano Shimoshinjo, 010-0195, Japan.
authors.forename: Akira
authors.initials: A
authors.lastname: Sasaki
authors.affiliation: Department of Biotechnology, Akita Prefectural University, 241-438 Kaidobata-Nishi, Akita City, Nakano Shimoshinjo, 010-0195, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Matsuo
authors.affiliation: Department of Biotechnology, Akita Prefectural University, 241-438 Kaidobata-Nishi, Akita City, Nakano Shimoshinjo, 010-0195, Japan.
authors.forename: Chika
authors.initials: C
authors.lastname: Yamamoto
authors.affiliation: Department of Biotechnology, Akita Prefectural University, 241-438 Kaidobata-Nishi, Akita City, Nakano Shimoshinjo, 010-0195, Japan.
authors.forename: Yuiko
authors.initials: Y
authors.lastname: Minato
authors.affiliation: Department of Biotechnology, Akita Prefectural University, 241-438 Kaidobata-Nishi, Akita City, Nakano Shimoshinjo, 010-0195, Japan.
authors.forename: Naoya
authors.initials: N
authors.lastname: Kuwahara
authors.affiliation: Department of Biotechnology, Akita Prefectural University, 241-438 Kaidobata-Nishi, Akita City, Nakano Shimoshinjo, 010-0195, Japan.
authors.forename: Chikako
authors.initials: C
authors.lastname: Fujii
authors.affiliation: Department of Biotechnology, Akita Prefectural University, 241-438 Kaidobata-Nishi, Akita City, Nakano Shimoshinjo, 010-0195, Japan.
authors.forename: Masayuki
authors.initials: M
authors.lastname: Kobayashi
authors.affiliation: Department of Applied Chemistry, National Institute of Technology, Akita College, Akita, 011-8511, Japan.
authors.forename: Hitosi
authors.initials: H
authors.lastname: Agematu
authors.affiliation: Bioproduction Research Institute, National Institute of Advanced Industrial Science and Technology (AIST), Sapporo, 062-8517, Japan.
authors.forename: Tomohiro
authors.initials: T
authors.lastname: Tamura
authors.affiliation: Laboratory of Microbial Engineering, Kitasato Institute for Life Sciences, Kitasato University, Sagamihara, Kanagawa, 252-0373, Japan.
authors.forename: Mamoru
authors.initials: M
authors.lastname: Komatsu
authors.affiliation: Department of Bioactive Molecules, National Institute of Infectious Diseases, Tokyo, 162-8640, Japan.
authors.forename: Jun
authors.initials: J
authors.lastname: Ishikawa
authors.affiliation: Laboratory of Microbial Engineering, Kitasato Institute for Life Sciences, Kitasato University, Sagamihara, Kanagawa, 252-0373, Japan.
authors.forename: Haruo
authors.initials: H
authors.lastname: Ikeda
authors.affiliation: Department of Biotechnology, Akita Prefectural University, 241-438 Kaidobata-Nishi, Akita City, Nakano Shimoshinjo, 010-0195, Japan.
authors.forename: Ikuo
authors.initials: I
authors.lastname: Kojima
mesh_terms: D000617:Aminoglycosides / Q000096:biosynthesis*; D000900:Anti-Bacterial Agents / Q000096:biosynthesis*; D001483:Base Sequence; D003001:Cloning, Molecular; D005285:Fermentation; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D007294:Inositol / Q000096:biosynthesis / Q000378:metabolism; D005810:Multigene Family; D007296:Myo-Inositol-1-Phosphate Synthase / Q000235:genetics / Q000378:metabolism; D012240:Rhodococcus / Q000235:genetics* / Q000378:metabolism; D013302:Streptomyces / Q000235:genetics* / Q000378:metabolism; D048170:Streptomyces lividans / Q000235:genetics*; D014157:Transcription Factors / Q000378:metabolism
chemical_list: D000617:Aminoglycosides; D000900:Anti-Bacterial Agents; D014157:Transcription Factors; D007294:Inositol; D007296:Myo-Inositol-1-Phosphate Synthase; C100305:kasugamycin
keywords: Heterologous production; Kasugamycin; Rhodococcus erythropolis; Streptomyces lividans