Identification of a Novel Staphylococcus aureus
Two-Component Leukotoxin Using Cell Surface Proteomics |
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Authors: | Christy L Ventura Natalia Malachowa Carl H Hammer Glenn A Nardone Mary Ann Robinson Scott D Kobayashi Frank R DeLeo |
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Institution: | 1. Laboratory of Human Bacterial Pathogenesis, Rocky Mountain Laboratories,
National Institute of Allergy and Infectious Diseases, National Institutes of
Health, Hamilton, Montana, United States of America.; 2. Research Technologies Branch, National Institute of Allergy and
Infectious Diseases, National Institutes of Health, Rockville, Maryland, United
States of America.;University of Liverpool, United Kingdom |
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Abstract: | Staphylococcus aureus is a prominent human pathogen and leading
cause of bacterial infection in hospitals and the community.
Community-associated methicillin-resistant S. aureus (CA-MRSA)
strains such as USA300 are highly virulent and, unlike hospital strains, often
cause disease in otherwise healthy individuals. The enhanced virulence of
CA-MRSA is based in part on increased ability to produce high levels of secreted
molecules that facilitate evasion of the innate immune response. Although
progress has been made, the factors that contribute to CA-MRSA virulence are
incompletely defined. We analyzed the cell surface proteome (surfome) of USA300
strain LAC to better understand extracellular factors that contribute to the
enhanced virulence phenotype. A total of 113 identified proteins were associated
with the surface of USA300 during the late-exponential phase of growth
in vitro. Protein A was the most abundant surface molecule
of USA300, as indicated by combined Mascot score following analysis of peptides
by tandem mass spectrometry. Unexpectedly, we identified a previously
uncharacterized two-component leukotoxin–herein named LukS-H and
LukF-G (LukGH)-as two of the most abundant surface-associated proteins of
USA300. Rabbit antibody specific for LukG indicated it was also freely secreted
by USA300 into culture media. We used wild-type and isogenic
lukGH deletion strains of USA300 in combination with human
PMN pore formation and lysis assays to identify this molecule as a leukotoxin.
Moreover, LukGH synergized with PVL to enhance lysis of human PMNs in
vitro, and contributed to lysis of PMNs after phagocytosis. We
conclude LukGH is a novel two-component leukotoxin with cytolytic activity
toward neutrophils, and thus potentially contributes to S.
aureus virulence. |
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