caa a22 4500 46575855X CHVBK 20180323111854.0 cr unu---uuuuu 170327e19900301xx s 000 0 eng 10.1007/BF00312612 doi (NATIONALLICENCE)springer-10.1007/BF00312612 Cloning and expression of a hybrid Streptomyces clavuligerus cef E gene in Penicillium chrysogenum [Elektronische Daten] [Cathleen Cantwell, Robert Beckmann, Joe Dotzlaf, Deborah Fisher, Paul Skatrud, Wu-Kuang Yeh, Stephen Queener] Summary: A hybrid cefE gene was constructed by juxtaposing promoter sequences from the Penicillium chrysogenum pcbC gene to the open reading frame of the Streptomyces clavuligerus cefE gene. In S. clavuligerus the cefE gene codes for the enzyme penicillin N expandase [also known as deacetoxycephalosporin C synthetase (DAOCS)]. To insert the hybrid cefE gene into P. chrysogenum the vector pPS65 was constructed; pPS65 contains the hybrid cefE gene and the Aspergillus nidulans amdS gene. The amdS gene encodes acetamidase and provides for dominant selection in P. chrysogenum. Protoplasts of P. chrysogenum were transformed with pPS65 and selected for the ability to grow on acetamide medium. Extracts of cells cultivated in penicillin production medium were analyzed for penicillin N expandase activity. Penicillin N expandase activity was detected in approximately one-third of the transformants tested. Transformants WG9-69C-01 and WG9-61L-03 had the highest specific activities of penicillin N expandase: 4.3% and 10.3%, respectively, relative to the amount of penicillin N expandase in S. clavuligerus. Untrasformed P. chrysogenum exhibited no penicillin N expandase activity. Analysis of the penicillin V titer revealed that WG9-61L-03 produced titers equal to that of the recipient strain while the amount of penicillin V produced in WG9-69C-01 was reduced by five fold. Springer-Verlag, 1990 Penicillium chrysogenum nationallicence Penicillin N expandase nationallicence Beta-lactam antibiotics nationallicence Heterologous gene expression nationallicence Cantwell Cathleen Lilly Research Laboratories, 46285, Indianapolis, IN, USA aut Beckmann Robert Lilly Research Laboratories, 46285, Indianapolis, IN, USA aut Dotzlaf Joe Lilly Research Laboratories, 46285, Indianapolis, IN, USA aut Fisher Deborah Lilly Research Laboratories, 46285, Indianapolis, IN, USA aut Skatrud Paul Lilly Research Laboratories, 46285, Indianapolis, IN, USA aut Yeh Wu-Kuang Lilly Research Laboratories, 46285, Indianapolis, IN, USA aut Queener Stephen Lilly Research Laboratories, 46285, Indianapolis, IN, USA aut Current Genetics Springer-Verlag 17/3(1990-03-01), 213-221 0172-8083 17:3<213 1990 17 294 https://doi.org/10.1007/BF00312612 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/BF00312612 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Cantwell Cathleen Lilly Research Laboratories, 46285, Indianapolis, IN, USA aut NATIONALLICENCE 700 1- Beckmann Robert Lilly Research Laboratories, 46285, Indianapolis, IN, USA aut NATIONALLICENCE 700 1- Dotzlaf Joe Lilly Research Laboratories, 46285, Indianapolis, IN, USA aut NATIONALLICENCE 700 1- Fisher Deborah Lilly Research Laboratories, 46285, Indianapolis, IN, USA aut NATIONALLICENCE 700 1- Skatrud Paul Lilly Research Laboratories, 46285, Indianapolis, IN, USA aut NATIONALLICENCE 700 1- Yeh Wu-Kuang Lilly Research Laboratories, 46285, Indianapolis, IN, USA aut NATIONALLICENCE 700 1- Queener Stephen Lilly Research Laboratories, 46285, Indianapolis, IN, USA aut NATIONALLICENCE 773 0- Current Genetics Springer-Verlag 17/3(1990-03-01), 213-221 0172-8083 17:3<213 1990 17 294 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer