Haemophilus ducreyi LspA proteins are tyrosine phosphorylated by macrophage-encoded protein tyrosine kinases

Kaiping Deng, Jason R. Mock, Steven Greenberg, Nicolai S C Van Oers, Eric J. Hansen

Research output: Contribution to journalArticle

9 Citations (Scopus)

Abstract

The LspA proteins (LspA1 and LspA2) of Haemophilus ducreyi are necessary for this pathogen to inhibit the phagocytic activity of macrophage cell lines, an event that can be correlated with a reduction in the level of active Src family protein tyrosine kinases (PTKs) in these eukaryotic cells. During studies investigating this inhibitory mechanism, it was discovered that the LspA proteins themselves were tyrosine phosphorylated after wild-type H. ducreyi cells were incubated with macrophages. LspA proteins in cell-free concentrated H. ducreyi culture supernatant fluid could also be tyrosine phosphorylated by macrophages. This ability to tyrosine phosphorylate the LspA proteins was not limited to immune cell lineages but could be accomplished by both HeLa and COS-7 cells. Kinase inhibitor studies with macrophages demonstrated that the Src family PTKs were required for this tyrosine phosphorylation activity. In silico methods and site-directed mutagenesis were used to identify EPIYG and EPVYA motifs in LspA1 that contained tyrosines that were targets for phosphorylation. A total of four tyrosines could be phosphorylated in LspA1, with LspA2 containing eight predicted tyrosine phosphorylation motifs. Purified LspA1 fusion proteins containing either the EPIYG or EPVYA motifs were shown to be phosphorylated by purified Src PTK in vitro. Macrophage lysates could also tyrosine phosphorylate the LspA proteins and an LspA1 fusion protein via a mechanism that was dependent on the presence of both divalent cations and ATP. Several motifs known to interact with or otherwise affect eukaryotic kinases were identified in the LspA proteins.

Original languageEnglish (US)
Pages (from-to)4692-4702
Number of pages11
JournalInfection and Immunity
Volume76
Issue number10
DOIs
StatePublished - Oct 2008

Fingerprint

Haemophilus ducreyi
Protein-Tyrosine Kinases
Tyrosine
Macrophages
src-Family Kinases
Proteins
Phosphorylation
Phosphotransferases
COS Cells
Divalent Cations
Eukaryotic Cells
Cell Lineage
Site-Directed Mutagenesis
Computer Simulation
Adenosine Triphosphate
Cell Line

ASJC Scopus subject areas

  • Immunology
  • Microbiology
  • Parasitology
  • Infectious Diseases

Cite this

Haemophilus ducreyi LspA proteins are tyrosine phosphorylated by macrophage-encoded protein tyrosine kinases. / Deng, Kaiping; Mock, Jason R.; Greenberg, Steven; Van Oers, Nicolai S C; Hansen, Eric J.

In: Infection and Immunity, Vol. 76, No. 10, 10.2008, p. 4692-4702.

Research output: Contribution to journalArticle

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