Cleavage of Tumor Necrosis Factor-alpha by Legionella Exoprotease

The role of the major secretory protein of Legionella pneumophila, a zinc protease, in Legionella infection is not known. Since an important step of the host reaction in Legionnaires' disease is the production of tumor necrosis factor-alpha (TNF-alpha) by alveolar macrophages, we studied the interaction of Legionella protease and U-937 cells with respect to TNF-alpha. The Legionella protease was purified by fractionated precipitation, gel filtration and hydrophobic interaction chromatography. The purified enzyme was added to U-937 cells, a promyelocytic cell line. In the supernatants of PMA-treated U-937 cells we found low concentrations of TNF-alpha after incubation with protease. Therefore we pursued the hypothesis of direct enzymatic degradation of TNF-alpha by Legionella protease. Enzymatic cleavage of TNF-alpha was proven by SDS-PAGE, ELISA and TNF-alpha bioassay with L-929 cells. The degradation of TNF-alpha by the Legionella protease was shown in all three systems. Enzymatic degradation of TNF-alpha might be important for the pathogenesis of Legionnaires' disease.