Isolation and characterization of hemolysin activated by reductant from Prevotella intermedia

• Published in: FEMS immunology and medical microbiology Vol. 35; no. 1; pp. 43 - 47
• Main Authors: Takada, Kazuko, Fukatsu, Akira, Otake, Shigeo, Hirasawa, Masatomo
• Format: Journal Article
• Language: English
• Published: Oxford, UK Blackwell Publishing Ltd 01.01.2003; Blackwell; Oxford University Press
• Subjects: Ammonium; Ammonium sulfate; Bacteriology; Biological and medical sciences; Calcium; Calcium ions; Cell culture; Culture Media; Dithiothreitol; Ethylenediaminetetraacetic acids; Fundamental and applied biological sciences. Psychology; Glutathione; Heat exchange; Heat treatment; Hemolysin; Hemolysin Proteins - biosynthesis; Hemolysin Proteins - drug effects; Hemolysin Proteins - isolation & purification; Hot Temperature; Hydrogen-Ion Concentration; Ion exchange; Magnesium; Microbiology; Oxidation; Pathogenicity, virulence, toxins, bacteriocins, pyrogens, host-bacteria relations, miscellaneous strains; Polyethylene glycol; Prevotella intermedia; Prevotella intermedia - growth & development; Prevotella intermedia - metabolism; Purification; Reducing Agents - pharmacology; Reductant; Trypsin; Trypsin - metabolism; Hemolysin; Reductant; Reduction; Purification; Prevotella intermedia; Bacteria; Chemical properties; Hemolysis test; Bacteroidaceae
• ISSN: 0928-8244; 1574-695X; 2049-632X
• DOI: 10.1111/j.1574-695X.2003.tb00647.x

Abstract The hemolysin from Prevotella intermedia was partially purified from culture supernatant and then characterized. The hemolysin produced a clear β-hemolytic zone on a blood agar plate. Hemolytic activity was 2.5-fold greater in culture supernatant compared to that cell-associated. The isolation and purification procedure involved ammonium sulfate and polyethylene glycol precipitations and ion-exchange chromatographies on DEAE-Sephacel and CM-Sepharose. The activity of this hemolysin was stimulated by reductants such as cysteine, dithiothreitol, glutathione etc., and was lost upon oxidation. Trypsin or heat treatment resulted in complete inhibition of hemolytic activity. Ca2+, Mg2+ and EDTA did not affect the activity. The optimal pH of this hemolysin was 7.5.