Abstract: Abstract: To ascertain the nature of gelatinase and its role in the pathogenesis of R. anatipestifer, the enzyme from R. anatipestifer strain AF, was purified and its characterizations were analyzed, including stability for heat, effects of proteases inhibitors and metal ions on its activities and pathogenicity for ducklings. The results revealed that gelatinase was purified from the culture supernatant of R. anatipestifer strain AF by ultrafiltration, 70% ammonium sulfate precipitation, anion exchange chromatography, hydrophobic interaction chromatography and gel permeation chromatography. The sample migrated as a single band on SDS- polyacrylamide gel electrophoresis with a molecular mass of 59000. Its activity was inhibited by EDTA, Cu2+, Cd2+, Mn2+, Hg2+, PMSF, NBS, AAD, and by heating at 65 ℃ for 6 min. Purified gelatinase was proved to be a direct virulent factor on the basis of eliciting lethal toxicity to duckling by intravenous inoculation. The gelatinase not only enhanced the invasion of R. anatipestifer, increased bacteremia, shortened stage of latency and course of disease, but also raised mortality obviously. The results indicated that the gelatinase is metalloprotease, and it is virulence important role in pathogenesis of R. anatipestifer.

Key words: Key words: Riemerella anatipestifer; gelatinase; purification; characterization