%0 Journal Article %1 Oey2009a %A Oey, M. %A Lohse, M. %A Scharff, L. B. %A Kreikemeyer, B. %A Bock, R. %D 2009 %J Proc Natl Acad Sci U S A %K Agents/*biosynthesis/*therapeutic Anti-Bacterial Antimicrobial Bacteriolysis/drug Biological/toxicity Biotechnology/*methods Cationic Data Genetic Genetic/drug Genome, Messenger/genetics/metabolism Microbial Molecular Peptides/*biosynthesis Plastid/genetics Plastids/*metabolism Pneumonia/*drug RNA, Sensitivity Sequence Streptococcus Tests Tobacco Toxins, Transcription, Transformation, Vectors/genetics Viability/drug effects pneumoniae/drug therapy use/toxicity %N 16 %P 6579-84 %T Plastid production of protein antibiotics against pneumonia via a new strategy for high-level expression of antimicrobial proteins %U http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19332784 %V 106 %X Plastid transformation has become an attractive tool in biotechnology. Because of the prokaryotic nature of the plastid's gene expression machinery, expression elements (promoters and untranslated regions) that trigger high-level foreign protein accumulation in plastids usually also confer high expression in bacterial cloning hosts. This can cause problems, for example, when production of antimicrobial compounds is attempted. Their bactericidal activity can make the cloning of the corresponding genes in plastid transformation vectors impossible. Here, we report a general solution to this problem. We have designed a strategy (referred to as toxin shuttle) that allows the expression in plastids of proteins that are toxic to Escherichia coli. The strategy is based on blocking transcription in E. coli by bacterial transcription terminators upstream of the gene of interest, which subsequently are excised in planta by site-specific recombination. We demonstrate the applicability of the strategy by the high-level expression in plastids (to up to 30% of the plant's total soluble protein) of 2 phage-derived protein antibiotics that are toxic to E. coli. We also show that the plastid-produced antibiotics efficiently kill pathogenic strains of Streptococcus pneumoniae, the causative agent of pneumonia, thus providing a promising strategy for the production of next-generation antibiotics in plants.

@article{Oey2009a, abstract = {Plastid transformation has become an attractive tool in biotechnology. Because of the prokaryotic nature of the plastid's gene expression machinery, expression elements (promoters and untranslated regions) that trigger high-level foreign protein accumulation in plastids usually also confer high expression in bacterial cloning hosts. This can cause problems, for example, when production of antimicrobial compounds is attempted. Their bactericidal activity can make the cloning of the corresponding genes in plastid transformation vectors impossible. Here, we report a general solution to this problem. We have designed a strategy (referred to as toxin shuttle) that allows the expression in plastids of proteins that are toxic to Escherichia coli. The strategy is based on blocking transcription in E. coli by bacterial transcription terminators upstream of the gene of interest, which subsequently are excised in planta by site-specific recombination. We demonstrate the applicability of the strategy by the high-level expression in plastids (to up to 30% of the plant's total soluble protein) of 2 phage-derived protein antibiotics that are toxic to E. coli. We also show that the plastid-produced antibiotics efficiently kill pathogenic strains of Streptococcus pneumoniae, the causative agent of pneumonia, thus providing a promising strategy for the production of next-generation antibiotics in plants.}, added-at = {2010-12-14T18:12:02.000+0100}, author = {Oey, M. and Lohse, M. and Scharff, L. B. and Kreikemeyer, B. and Bock, R.}, biburl = {https://www.bibsonomy.org/bibtex/201efbe94a272d178360b67af75ff8cab/pharmawuerz}, endnotereftype = {Journal Article}, interhash = {41b3fd42d446a4f91e6e396e0eb7c889}, intrahash = {01efbe94a272d178360b67af75ff8cab}, issn = {1091-6490 (Electronic) 1091-6490 (Linking)}, journal = {Proc Natl Acad Sci U S A}, keywords = {Agents/*biosynthesis/*therapeutic Anti-Bacterial Antimicrobial Bacteriolysis/drug Biological/toxicity Biotechnology/*methods Cationic Data Genetic Genetic/drug Genome, Messenger/genetics/metabolism Microbial Molecular Peptides/*biosynthesis Plastid/genetics Plastids/*metabolism Pneumonia/*drug RNA, Sensitivity Sequence Streptococcus Tests Tobacco Toxins, Transcription, Transformation, Vectors/genetics Viability/drug effects pneumoniae/drug therapy use/toxicity}, month = {Apr 21}, note = {Oey, Melanie Lohse, Marc Scharff, Lars B Kreikemeyer, Bernd Bock, Ralph Research Support, Non-U.S. Gov't United States Proceedings of the National Academy of Sciences of the United States of America Proc Natl Acad Sci U S A. 2009 Apr 21;106(16):6579-84. Epub 2009 Mar 30.}, number = 16, pages = {6579-84}, shorttitle = {Plastid production of protein antibiotics against pneumonia via a new strategy for high-level expression of antimicrobial proteins}, timestamp = {2010-12-14T18:12:27.000+0100}, title = {Plastid production of protein antibiotics against pneumonia via a new strategy for high-level expression of antimicrobial proteins}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19332784}, volume = 106, year = 2009 }