Purification and characterization of hemolysin from Porphyromonas gingivalis A7436.

Porphyromonas gingivalis, a periodontal pathogen, has the ability to lyse erythrocytes. The hemolytic activity of P. gingivalis A7436 was purified as a 45-kDa protein from the culture supernatant of a 3-days old culture using nickel-nitrilotriacetic acid chromatography. Erythrocytes treated with purified P. gingivalis hemolysin showed the presence of pores and extracellular debris by scanning electron microscopy. Active immunization of mice with 15 micrograms hemolysin induced neutralizing antibodies to hemolysin. Heating at 60 degrees C and treatment with trypsin and dithiothreitol abolished hemolytic activity, while incubation with the protease inhibitor Na-p-tosyl-L-lysine chloromethyl ketone caused no effect. We report here for the first time purification of a hemolysin from P. gingivalis A7436. The amino acid sequence of an internal peptide of hemolysin showed sequence similarity with fimbrillin from P. gingivalis HG564. However, the amino acid composition of purified hemolysin was different from that of P. gingivalis fimbrillin. Also, the ability to lyse but not agglutinate erythrocytes and to bind to nickel-nitrilotriacetic acid differentiates P. gingivalis hemolysin from fimbrillin.