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Systematic analysis of the kalimantacin assembly line NRPS module using an adapted targeted mutagenesis approach
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Uytterhoeven, Birgit, Appermans, Kenny, Song, Lijiang, Masschelein, Joleen, Lathouwers, Thomas , Michiels, Chris W. and Lavigne, Rob (2016) Systematic analysis of the kalimantacin assembly line NRPS module using an adapted targeted mutagenesis approach. Microbiology Open, 5 (2). pp. 279-286. doi:10.1002/mbo3.326 ISSN 2045-8827.
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Official URL: http://doi.org/10.1002/mbo3.326
Abstract
Kalimantacin is an antimicrobial compound with strong antistaphylococcal activity that is produced by a hybrid trans-acyltransferase polyketide synthase/nonribosomal peptide synthetase system in Pseudomonas fluorescens BCCM_ID9359. We here present a systematic analysis of the substrate specificity of the glycine-incorporating adenylation domain from the kalimantacin biosynthetic assembly line by a targeted mutagenesis approach. The specificity-conferring code was adapted for use in Pseudomonas and mutated adenylation domain active site sequences were introduced in the kalimantacin gene cluster, using a newly adapted ligation independent cloning method. Antimicrobial activity screens and LC-MS analyses revealed that the production of the kalimantacin analogues in the mutated strains was abolished. These results support the idea that further insight in the specificity of downstream domains in nonribosomal peptide synthetases and polyketide synthases is required to efficiently engineer these strains in vivo.
| Item Type: | Journal Article | ||||||||
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| Subjects: | Q Science > QD Chemistry Q Science > QR Microbiology |
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| Divisions: | Faculty of Science, Engineering and Medicine > Science > Chemistry | ||||||||
| Library of Congress Subject Headings (LCSH): | Metabolites, Pseudomonas fluorescens, Mutagenesis, Polyketides, Ligases | ||||||||
| Journal or Publication Title: | Microbiology Open | ||||||||
| Publisher: | Wiley | ||||||||
| ISSN: | 2045-8827 | ||||||||
| Official Date: | April 2016 | ||||||||
| Dates: |
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| Volume: | 5 | ||||||||
| Number: | 2 | ||||||||
| Page Range: | pp. 279-286 | ||||||||
| DOI: | 10.1002/mbo3.326 | ||||||||
| Status: | Peer Reviewed | ||||||||
| Publication Status: | Published | ||||||||
| Access rights to Published version: | Restricted or Subscription Access | ||||||||
| Date of first compliant deposit: | 20 December 2017 | ||||||||
| Date of first compliant Open Access: | 21 December 2017 | ||||||||
| Open Access Version: |
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