title: Development of a high-expression system for staphylococcal exfoliative toxin genes.
abstract: We constructed a new expression system for staphylococcal exfoliative toxin (ET). The expression vector, pETA-exp2, was constructed based on Bacillus-Escherichia shuttle vector pHY300PLK. The pETA-exp2 vector includes the regulator of the ETA gene (eta), the promoter and Shine-Dalgarno (SD) sequences of eta, a SalI sequence at the end of the signal sequence of eta, a nucleotide sequence encoding mature ETA, an XhoI site, a 6x His sequence just before the stop codon and the end of the transcription sequence of eta. The nucleotide sequences coding for the mature proteins of ETB, ExhA, ExhB, ExhC, ExhD and SHETB were amplified by polymerase chain reaction (PCR) and inserted into pETA-exp2. These recombinant plasmids were transformed into Bacillus megaterium. The major protein in the culture supernatant of the transformant was recombinant ET (rET). The yields of all rETs were high, and all of them showed exfoliative activity in susceptible animals. The antigenicities of rETs and ETs were not distinguishable from each other.
authors.affiliation: Laboratory of Veterinary Microbiology, Faculty of Veterinary Medicine, School of Veterinary Medicine, Kitasato University, 23–35–1 Higashi, Towada, Aomori 034–8628, Japan.
authors.forename: Kenta
authors.initials: K
authors.lastname: Onuma
authors.forename: Taishi
authors.initials: T
authors.lastname: Tanabe
authors.forename: Hisaaki
authors.initials: H
authors.lastname: Sato
mesh_terms: D000818:Animals; D001410:Bacillus megaterium / Q000235:genetics*; D015153:Blotting, Western; D004269:DNA, Bacterial / Q000235:genetics; D004274:DNA, Recombinant; D004591:Electrophoresis, Polyacrylamide Gel; D005083:Exfoliatins / Q000096:biosynthesis* / Q000235:genetics / Q000633:toxicity; D005260:Female; D015964:Gene Expression Regulation, Bacterial; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D051379:Mice; D008807:Mice, Inbred BALB C; D010957:Plasmids; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins; D017422:Sequence Analysis, DNA; D057788:Staphylococcus hyicus / Q000235:genetics*; D013552:Swine; D014169:Transformation, Bacterial
chemical_list: D004269:DNA, Bacterial; D004274:DNA, Recombinant; D005083:Exfoliatins; D011994:Recombinant Proteins