Abstract
Legionella pneumophila is a Gram-negative pathogenic bacterium that causes severe pneumonia in humans. It establishes a replicative niche called Legionella-containing vacuole (LCV) that allows bacteria to survive and replicate inside pulmonary macrophages. To hijack host cell defense systems, L. pneumophila injects over 300 effector proteins into the host cell cytosol. The Lem4 effector (lpg1101) consists of two domains: an N-terminal haloacid dehalogenase (HAD) domain with unknown function and a C-terminal phosphatidylinositol 4-phosphate-binding domain that anchors Lem4 to the membrane of early LCVs. Herein, we demonstrate that the HAD domain (Lem4-N) is structurally similar to mouse MDP-1 phosphatase and displays phosphotyrosine phosphatase activity. Substrate specificity of Lem4 was probed using a tyrosine phosphatase substrate set, which contained a selection of 360 phosphopeptides derived from human phosphorylation sites. This assay allowed us to identify a consensus pTyr-containing motif. Based on the localization of Lem4 to lysosomes and to some extent to plasma membrane when expressed in human cells, we hypothesize that this protein is involved in protein-protein interactions with an LCV or plasma membrane-associated tyrosine-phosphorylated host target.
Highlights
Legionella pneumophila is a Gram-negative pathogenic bacterium that causes severe pneumonia in humans
The Lem4-N fold belongs to the large haloacid dehalogenase (HAD)-like protein superfamily, containing enzymatic activities such as phosphatases, phosphonatases, P-type ATPases, phosphoglucomutases, phosphomannomutases, and dehalogenases, which are all involved in a variety of cellular processes ranging from amino acid biosynthesis to detoxification [26]
Secreted effectors of L. pneumophila were shown to interfere with mitogen-activated protein kinase (MAPK) [40] and NF-B pathways to promote infection [41]
Summary
Legionella pneumophila is a Gram-negative pathogenic bacterium that causes severe pneumonia in humans. It establishes a replicative niche called Legionella-containing vacuole (LCV) that allows bacteria to survive and replicate inside pulmonary macrophages. Substrate specificity of Lem was probed using a tyrosine phosphatase substrate set, which contained a selection of 360 phosphopeptides derived from human phosphorylation sites. This assay allowed us to identify a consensus pTyr-containing motif. Based on the localization of Lem to lysosomes and to some extent to plasma membrane when expressed in human cells, we hypothesize that this protein is involved in protein–protein interactions with an LCV or plasma membrane–associated tyrosine-phosphorylated host target. The atomic coordinates and structure factors (codes 6CGJ, 6CGK, and 6CDW)
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