title: Immunoaffinity purification, stabilization and comparative characterization of listeriolysin O from Listeria monocytogenes serotypes 1/2a and 4b.
abstract: We developed a simple and highly effective procedure for stabilizing the haemolytic activity of listeriolysin O (LLO) from Listeria monocytogenes after immunoaffinity purification. The haemolytic activity of LLO was stabilized by eluting it directly into tubes containing an alkaline buffer (5 mM lysine, 140 mM KCl, 50% ethylene glycol, pH 11.5). The purified LLO retained 100% of its haemolytic activity after 6 weeks of storage at -20 degrees C. LLO purified from a strain of L. monocytogenes serotype 1/2a (ATCC 43249) and LLO purified from a strain of L. monocytogenes serotype 4b (F 2365) isolated from a Mexican-style cheese, showed no significant differences in pH and temperature stability. When incubated in buffers at pH values from 4 to 12 at 4 degrees C and 25 degrees C, LLO from serotypes 1/2a and 4b retained maximal haemolytic activity at pH 8 after 4 h of incubation. LLO from both serotypes lost their haemolytic activity after incubation at 50 degrees C for 25 min.
authors.affiliation: Division of Bacterial and Mycotic Diseases, Centers for Disease Control, Atlanta, GA 30333.
authors.forename: G M
authors.initials: GM
authors.lastname: Matar
authors.forename: W F
authors.initials: WF
authors.lastname: Bibb
authors.forename: L
authors.initials: L
authors.lastname: Helsel
authors.forename: W
authors.initials: W
authors.lastname: Dewitt
authors.forename: B
authors.initials: B
authors.lastname: Swaminathan
mesh_terms: D001427:Bacterial Toxins; D002846:Chromatography, Affinity / Q000379:methods*; D004355:Drug Stability; D006360:Heat-Shock Proteins / Q000737:chemistry / Q000302:isolation & purification*; D006460:Hemolysin Proteins; D006863:Hydrogen-Ion Concentration; D015151:Immunoblotting; D066298:In Vitro Techniques; D008089:Listeria monocytogenes / Q000378:metabolism* / Q000472:pathogenicity; D014774:Virulence
chemical_list: D001427:Bacterial Toxins; D006360:Heat-Shock Proteins; D006460:Hemolysin Proteins; C038705:hlyA protein, Listeria monocytogenes