TY  - JOUR
TI  - easyPACId, a Simple Method for Induced Production, Isolation, Identification, and Testing of Natural Products from Proteobacteria
AU  - Bode, Edna 
AU  - Assmann, Daniela 
AU  - Happel, Petra 
AU  - Meyer, Elmar 
AU  - Münch, Karin 
AU  - Rössel, Nicole 
AU  - Bode, Helge B.
VL  - 13
IS  - 13
PY  - 2023
DA  - 2023/07/05
SP  - e4709
C1  - Bio-protocol 2023;13:e4709
DO  - 10.21769/BioProtoc.4709
UR  - https://doi.org/10.21769/BioProtoc.4709
AB  - The easyPACId (easy Promoter Activation and Compound Identification) approach is focused on the targeted activation of natural product biosynthetic gene clusters (BGCs) encoding non-ribosomal peptide synthetases (NRPS), polyketide synthases (PKS), NRPS-PKS hybrids, or other BGC classes. It was applied to entomopathogenic bacteria of the genera Xenorhabdus and Photorhabdus by exchanging the natural promoter of desired BGCs against the L-arabinose inducible PBAD promoter in hfq mutants of the respective strains. The crude (culture) extracts of the cultivated easyPACId mutants are enriched with the single compound or compound class and can be tested directly against various target organisms without further purification of the produced natural products. Furthermore, isolation and identification of compounds from these mutants is simplified due to the reduced background in the hfq strains. The approach avoids problems often encountered in heterologous expression hosts, chemical synthesis, or tedious extraction of desired compounds from wild-type crude extracts. This protocol describes easyPACId for Xenorhabdus and Photorhabdus, but it was also successfully adapted to Pseudomonas entomophila and might be suitable for other proteobacteria that carry hfq.
KW  - easyPACId
KW  - Biosynthetic gene cluster activation
KW  - NRPS
KW  - PKS
KW  - Natural product
KW  - Promoter exchange
KW  - Bioactivity testing
KW  - Bioactive compounds isolation
KW  - Xenorhabdus
KW  - Photorhabdus
KW  - Pseudomonas
JF  - Bio-protocol
SN  - 2331-8325
PB  - Bio-protocol LLC.
BIO101  - False