Separation and properties of the haemolysins and extracellular enzymes of Listeria monocytogenes and L. ivanovii.
Desalted ammonium-sulphate (0-65%) precipitates from the cell-free supernates of 16-24-h cultures of Listeria monocytogenes Boldy and L. ivanovii (previously L. monocytogenes) Type 5 were eluted through Sephadex G-200. The enzyme activities gave rise to two main peaks. The first peak (approximate mol. wt of protein 150,000) contained only phosphatase activity (assayed by hydrolysis of 4-nitrophenylphosphate at pH 5.0 and 7.0). The second peak (approximate mol. wts of proteins 40,000-60,000) contained the haemolysin activity and the following hydrolytic activities (assay substrates are given in parentheses): phospholipase C (phosphatidyl choline and 4-nitrophenyl-phosphoryl-choline); phosphodiesterase (bis-4-nitrophenyl-phosphate); acid phosphatase (4-nitrophenylphosphatase); and esterases and lipases (4-nitrophenyl acetate, naphthyl-acetate and -oleate, triacetin and triolein). DEAE-Sephadex chromatography of appropriate fractions from the Sephadex G-200 purification step separated the first peak into two phosphatases and resolved the second peak into its constituent activities. Polyacrylamide gel electrophoresis showed that the individual fractions from the DEAE-Sephadex step consisted of mixtures of protein. The effects of pH and potential activators and inhibitors on the active proteins purified by DEAE-Sephadex chromatography were examined.
Barclay
R
R
Department of Plant Biology, University of Hull.
Threlfall
D R
DR
Leighton
I
I
D000907:Antibodies, Bacterial / Q000276:immunology; D002413:Cations, Divalent / Q000494:pharmacology; D002784:Cholesterol / Q000494:pharmacology; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D006460:Hemolysin Proteins / Q000302:isolation & purification*; D008087:Listeria / Q000032:analysis*; D008089:Listeria monocytogenes / Q000032:analysis*; D010710:Phosphates / Q000494:pharmacology; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology; D010744:Phosphoric Monoester Hydrolases / Q000302:isolation & purification* / Q000378:metabolism; D010738:Type C Phospholipases / Q000378:metabolism; D015032:Zinc / Q000494:pharmacology
D000907:Antibodies, Bacterial; D002413:Cations, Divalent; D006460:Hemolysin Proteins; D010710:Phosphates; D010726:Phosphodiesterase Inhibitors; D002784:Cholesterol; D010744:Phosphoric Monoester Hydrolases; D010738:Type C Phospholipases; D015032:Zinc