Proteomics-Based Identification of Anchorless Cell Wall Proteins as Vaccine Candidates againstStaphylococcus aureus

Abstract

Staphylococcus aureus is an important human pathogen with increasing clinical impact due to the extensive spread of antibiotic-resistant strains. Therefore, development of a protective polyvalent vaccine is of great clinical interest. We employed an intravenous immunoglobulin (IVIG) preparation as a source of antibodies directed against anchorless S. aureus surface proteins for identification of novel vaccine candidates. In order to identify such proteins, subtractive proteome analysis (SUPRA) of S. aureus anchorless cell wall proteins was performed. Proteins reacting with IVIG but not with IVIG depleted of S. aureus-specific opsonizing antibodies were considered vaccine candidates. Nearly 40 proteins were identified by this preselection method using matrix-assisted laser desorption ionization-time of flight analysis. Three of these candidate proteins, enolase (Eno), oxoacyl reductase (Oxo), and hypothetical protein hp2160, were expressed as glutathione S-transferase fusion proteins, purified, and used for enrichment of corresponding immunoglobulin Gs from IVIG by affinity chromatography. Use of affinity-purified anti-Eno, anti-Oxo, and anti-hp2160 antibodies resulted in opsonization, phagocytosis, and killing of S. aureus by human neutrophils. High specific antibody titers were detected in mice immunized with recombinant antigens. In mice challenged with bioluminescent S. aureus, reduced staphylococcal spread was measured by in vivo imaging. The recovery of S. aureus CFU from organs of immunized mice was diminished 10-to 100-fold. Finally, mice immunized with hp2160 displayed statistically significant higher survival rates after lethal challenge with clinically relevant S. aureus strains. Taken together, our data suggest that anchorless cell wall proteins might be promising vaccine candidates and that SUPRA is a valuable tool for their identification.

Biofilm Matrix Exoproteins Induce a Protective Immune Response against Staphylococcus aureus Biofilm Infection

“…Thus, a total of 33 extracellular proteins were detected with a MASCOT score higher than 50 (Table 2). Importantly, the proteins identified have been repeatedly detected in extracellular proteomes of various S. aureus isolates (34,(47)(48)(49)(50)(51)(52)(53)(54). More notably, 28 out of the 33 proteins identified in our analysis have also been found in the biofilm exoproteome of S. aureus D30 strain, isolated from a persistent nasal carrier (Table 2) (50).…”

“…Success of S. aureus as a pathogen is the result of different abilities, such as the capacity to invade a wide variety of cell types, to secrete a diversity of proteins and toxins, and to persist in the host, remaining resistant to clearance by the immune system or antibiotics through a biofilm mode of growth. Numerous approaches have been adopted in order to identify staphylococcal surface-and cell wall-associated proteins as antigenic candidates for a vaccine against S. aureus infections (34,49,51,53,(56)(57)(58)(59)(60). However, few works have been focused on the selection of antigens that could also protect against biofilm-associated bacteria (14,(24)(25)(26).…”

Genomic and Surface Proteomic Analysis of the Canine Pathogen Staphylococcus pseudintermedius Reveals Proteins That Mediate Adherence to the Extracellular Matrix

“…The identification of intracellular proteins on the cell surface is not unexpected, and numerous studies of S. aureus have reported a similar phenomenon (10,16,46,53,58). In some cases, alternative functions associated with the cell surface have been described for cytoplasmic proteins, including GAPDH (26,52), enolase (4,23), transcription elongation factors (1,24), and chaperone proteins GroES and DnaK (20,57).…”

Biomedical Mass Spectrometry in Today's and Tomorrow's Clinical Microbiology Laboratories

“…When applying MS technologies, the amino acid sequences of specific target proteins can be determined (15). However, the diagnostic MALDI-TOF MS systems as marketed today are focused primarily on bacterial species identification.…”