title: Development of nitrilase promoter-derived inducible vectors for Streptomyces.
abstract: An inducible expression vector, pSH19, which harbors regulatory expression system PnitA-NitR, for streptomycetes was constructed previously. Here, we have modified pSH19 to obtain shuttle vectors for Streptomyces-E. coli by introducing the replication origin of a plasmid for E. coli (ColE1) and an antibiotic-resistant gene. Six inducible shuttle vectors, pESH19cF, pESH19cR, pESH19kF, pESH19kR, pESH19aF, and pESH19aR, for Streptomyces-E. coli, were successfully developed. The stability of these vectors was examined in five different E. coli strains and Streptomyces lividans TK24. The stability test showed that the pSH19-derived shuttle vectors were stable in E. coli Stbl2 and S. lividans TK24. Heterologous expression experiments involving each of the catechol 2,3-dioxygenase, nitrilase, and N-substituted formamide deformylase genes as a reporter gene showed that pESH19cF, pESH19kF, and pESH19aF possess inducible expression ability in S. lividans TK24. Thus, these vectors were found to be useful expression tools for experiments on both Gram-negative and Gram-positive bacterial genes.
authors.affiliation: a Institute of Applied Biochemistry, and Graduate School of Life and Environmental Sciences , The University of Tsukuba , Tsukuba , Japan.
authors.forename: Masako
authors.initials: M
authors.lastname: Matsumoto
authors.affiliation: a Institute of Applied Biochemistry, and Graduate School of Life and Environmental Sciences , The University of Tsukuba , Tsukuba , Japan.
authors.forename: Yoshiteru
authors.initials: Y
authors.lastname: Hashimoto
authors.affiliation: a Institute of Applied Biochemistry, and Graduate School of Life and Environmental Sciences , The University of Tsukuba , Tsukuba , Japan.
authors.forename: Yuki
authors.initials: Y
authors.lastname: Saitoh
authors.affiliation: a Institute of Applied Biochemistry, and Graduate School of Life and Environmental Sciences , The University of Tsukuba , Tsukuba , Japan.
authors.forename: Takuto
authors.initials: T
authors.lastname: Kumano
authors.affiliation: a Institute of Applied Biochemistry, and Graduate School of Life and Environmental Sciences , The University of Tsukuba , Tsukuba , Japan.
authors.forename: Michihiko
authors.initials: M
authors.lastname: Kobayashi
mesh_terms: D000581:Amidohydrolases / Q000235:genetics / Q000378:metabolism; D000619:Aminohydrolases / Q000235:genetics* / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D050557:Catechol 2,3-Dioxygenase / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D017930:Genes, Reporter; D005818:Genetic Engineering; D005822:Genetic Vectors / Q000737:chemistry / Q000378:metabolism*; D010957:Plasmids / Q000737:chemistry / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D048170:Streptomyces lividans / Q000235:genetics* / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D050557:Catechol 2,3-Dioxygenase; D000581:Amidohydrolases; C493455:N-substituted formamide deformylase; D000619:Aminohydrolases; C021867:nitrilase
keywords: Streptomyces; inducible expression; shuttle vector