Production of lipase by clinical isolates of Pseudomonas cepacia.
Ten clinical isolates of Pseudomonas cepacia from the sputum of cystic fibrosis patients were examined for the ability to produce lipase. Lipase substrates used included egg yolk agar, four different polyoxyethylene sorbitans (Tweens), and p-nitrophenylphosphorylcholine, a chromogenic substrate used to assay for phospholipase C. Lipase activity was detected in the filtrates of organisms grown to the exponential phase in either tryptose minimal medium or chemically defined medium. Lipase activity increased in the filtrates if the cultures were allowed to proceed into the stationary phase. None of the isolates produced phospholipase C. Lipase activity on Tween 20 ranged from 41.6 X 10(-3) to 640.0 X 10(-3) U/micrograms of protein. The activity was similar or slightly lower when Tween 40, 60, or 80 was used as the substrate. There was no correlation between lipase activity on Tween and that demonstrated on egg yolk agar. Lipase activity increased as pH increased from 7.0 to 9.0. Boiling for 5 min resulted in 66% loss of enzyme activity. The remaining activity continued to decrease with increasing boiling time. The enzyme was purified by gel filtration on Sephadex G-200, and the resultant preparation, when subjected to polyacrylamide gel electrophoresis, resulted in a single protein band (molecular weight, approximately 25,000) from which lipase activity could be eluted. The purified lipase was not cytotoxic to HeLa cells, nor was it toxic when injected intravenously into mice.
Lonon
M K
MK
Department of Microbiology, Texas Tech University Health Sciences Center, Lubbock 79430.
Woods
D E
DE
Straus
D C
DC
D000818:Animals; D002850:Chromatography, Gel; D003470:Culture Media; D003550:Cystic Fibrosis / Q000382:microbiology; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D006367:HeLa Cells; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008049:Lipase / Q000096:biosynthesis* / Q000633:toxicity; D051379:Mice; D011136:Polysorbates; D011549:Pseudomonas / Q000201:enzymology* / Q000254:growth & development; D013183:Sputum / Q000382:microbiology
D003470:Culture Media; D011136:Polysorbates; D008049:Lipase