Chapter 58 : Identification and Characterization of Legionella pneumophila Phospholipases A

Abstract:

The major proteins of the anion-exchange fractions with phospholipases A (PLA) activity were N-terminally sequenced. This chapter analyzes the bacterial membrane lipid composition of the Legionella pneumophila aas mutants and the 130b wild type by thin-layer chro-matography (TLC). The role of Aas during L. pneumophila intracellular infection was assessed in the three host models, U937 macrophages, A549 epithelial cells, and Acanthamoeba castellanii amoebae, and Aas was found to be dispensable, because L. pneumophila aas mutants showed the same increase in CFU in all three hosts during 72 h of infection as the wild type. The chapter presents data that show that L. pneumophila Aas contributes to the incorporation of phospholipids into the bacterial membrane, and in addition to L. pneumophila PlaA, is another enzyme involved in the detoxification of lysophospholipids. The authors examined the corresponding L. pneumophila 130b unk1 mutants for hydrolysis of diacylphospho-lipids, monoacylphospholipids, and 1-MPG and found that Unk1 did not contribute to the secreted PLA and L. pneumophila phospholipases A (LPLA) activities of L. pneumophila. The data indicate an essential role for Unk1 in infections of amoebae and macrophages by L. pneumophila. The authors identified three proteins, Aas, LvrE, and Unk1, present in the L. pneumophila culture supernatant. Aas contributes to the cell membrane integrity of L. pneumophila, but is dispensable for the infection of host cells. Since bacterial lipolytic enzymes are important bacterial tools for surviving both inside and outside of hosts, their characterization promotes one's understanding of the life cycle of L. pneumophila.