Inactivation of Lgt allows systematic characterization of lipoproteins from Listeria monocytogenes.
Lipoprotein anchoring in bacteria is mediated by the prolipoprotein diacylglyceryl transferase (Lgt), which catalyzes the transfer of a diacylglyceryl moiety to the prospective N-terminal cysteine of the mature lipoprotein. Deletion of the lgt gene in the gram-positive pathogen Listeria monocytogenes (i) impairs intracellular growth of the bacterium in different eukaryotic cell lines and (ii) leads to increased release of lipoproteins into the culture supernatant. Comparative extracellular proteome analyses of the EGDe wild-type strain and the Delta lgt mutant provided systematic insight into the relative expression of lipoproteins. Twenty-six of the 68 predicted lipoproteins were specifically released into the extracellular proteome of the Delta lgt strain, and this proved that deletion of lgt is an excellent approach for experimental verification of listerial lipoproteins. Consequently, we generated Delta lgt Delta prfA double mutants to detect lipoproteins belonging to the main virulence regulon that is controlled by PrfA. Overall, we identified three lipoproteins whose extracellular levels are regulated and one lipoprotein that is posttranslationally modified depending on PrfA. It is noteworthy that in contrast to previous studies of Escherichia coli, we unambiguously demonstrated that lipidation by Lgt is not a prerequisite for activity of the lipoprotein-specific signal peptidase II (Lsp) in Listeria.
Baumgärtner
Maja
M
Department of Cell Biology, Helmholtz Centre for Infection Research (HZI), D-38124 Braunschweig, Germany.
Kärst
Uwe
U
Gerstel
Birgit
B
Loessner
Martin
M
Wehland
Jürgen
J
Jänsch
Lothar
L
D016475:3T3 Cells; D000818:Animals; D016282:Aspartic Acid Endopeptidases / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D018938:Caco-2 Cells; D019295:Computational Biology; D015180:Electrophoresis, Gel, Two-Dimensional; D017353:Gene Deletion; D015964:Gene Expression Regulation, Bacterial; D006801:Humans; D008074:Lipoproteins / Q000378:metabolism*; D008089:Listeria monocytogenes / Q000235:genetics* / Q000378:metabolism / Q000472:pathogenicity; D051379:Mice; D008856:Microscopy, Fluorescence; D009154:Mutation; D009876:Operon / Q000235:genetics; D010454:Peptide Termination Factors / Q000235:genetics / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014166:Transferases / Q000235:genetics* / Q000378:metabolism; D014774:Virulence / Q000235:genetics
D001426:Bacterial Proteins; D008074:Lipoproteins; D010454:Peptide Termination Factors; D014166:Transferases; C042308:phosphatidylglycerol glyceryl transferase; D016282:Aspartic Acid Endopeptidases; C095652:signal peptidase II