Characterization of temperature-sensitive and lipopolysaccharide overproducing transposon mutants of Pseudomonas putida CA-3 affected in PHA accumulation.
A library of 20 000 transposon (Tn5) mutants of the gram-negative bacterium Pseudomonas putida CA-3 was generated and screened for adverse affects in polyhydroxyalkanoates (PHA) accumulation. Two mutants of interest were characterized phenotypically. CA-3-126, a mutant disrupted in a stress-related protein Clp protease subunit ClpA, demonstrated greater decreases in PHA accumulation compared with the wild type at reduced and elevated temperatures under PHA-accumulating growth conditions. CA-3-M, which is affected in the aminotransferase class I enzyme, accumulated reduced levels of PHA relative to the wild type and had lower growth yields on all carbon sources tested. Mutant CA-3-M produced up to 10-fold higher levels of lipopolysaccharide relative to the wild type and exhibited 1.2-fold lower aminotransferase activity with phenylalanine as a substrate compared with the wild-type strain. The composition of the lipopolysaccharide produced by the mutant differed from that produced by the wild-type strain. Growth and PHA accumulation by CA-3-M was the same as the wild type when the nitrogen concentration in the medium was increased to 265 mg N L(-1).
Goff
Miriam
M
Centre for Synthesis and Chemical Biology, Conway Institute for Biomolecular and Biomedical Research, University College Dublin, National University of Ireland, Belfield, Dublin, Ireland.
Nikodinovic-Runic
Jasmina
J
O'Connor
Kevin E
KE
D001426:Bacterial Proteins / Q000235:genetics; D018533:Biomass; D002244:Carbon / Q000378:metabolism; D004251:DNA Transposable Elements; D006358:Hot Temperature; D008070:Lipopolysaccharides / Q000378:metabolism*; D016254:Mutagenesis, Insertional; D009584:Nitrogen / Q000378:metabolism; D010447:Peptide Hydrolases / Q000235:genetics; D054813:Polyhydroxyalkanoates / Q000378:metabolism*; D016958:Pseudomonas putida / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism*; D000637:Transaminases / Q000235:genetics / Q000378:metabolism
D001426:Bacterial Proteins; D004251:DNA Transposable Elements; D008070:Lipopolysaccharides; D054813:Polyhydroxyalkanoates; D002244:Carbon; D000637:Transaminases; C022340:phenylalanine aminotransferase; D010447:Peptide Hydrolases; D009584:Nitrogen