Molecular mechanism of elongation factor 1A inhibition by a Legionella pneumophila glycosyltransferase

Ramon Hurtado-Guerrero*, Tal Zusman, Shalini Pathak, Adel F.M. Ibrahim, Sharon Shepherd, Alan Prescott, Gil Segal, Daan M.F. Van Aalten

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

Abstract

Legionnaires' disease is caused by a lethal colonization of alveolar macrophages with the Gram-negative bacterium Legionella pneumophila. LpGT (L. pneumophila glucosyltransferase; also known as Lgt1) has recently been identified as a virulence factor, shutting down protein synthesis in the human cell by specific glucosylation of EF1A (elongation factor 1A), using an unknown mode of substrate recognition and a retaining mechanism for glycosyl transfer. We have determined the crystal structure of LpGT in complex with substrates, revealing a GT-A fold with two unusual protruding domains. Through structure-guided mutagenesis of LpGT, several residues essential for binding of the UDP-glucose-donor and EF1A-acceptor substrates were identified, which also affected L. pneumophila virulence as demonstrated by microinjection studies. Together, these results suggested that a positively charged EF1A loop binds to a negatively charged conserved groove on the LpGTstructure, and that two asparagine residues are essential for catalysis. Furthermore, we showed that two further L. pneumophila glycosyltransferases possessed the conserved UDP-glucose-binding sites and EF1A-binding grooves, and are, like LpGT, translocated into the macrophage through the Icm/Dot (intracellular multiplication/defect in organelle trafficking) system.

Original languageEnglish
Pages (from-to)281-292
Number of pages12
JournalBiochemical Journal
Volume426
Issue number3
DOIs
StatePublished - 15 Mar 2010

Funding

FundersFunder number
Medical Research CouncilG0500367, G0900138

    Keywords

    • Elongation factor 1A (EF1A)
    • Glucosyl transferase
    • Legionella pneumophila
    • Microinjection
    • Protein structure
    • Site-directed mutagenesis

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