Functional selection of vaccine candidate peptides from Staphylococcus aureus whole-genome expression libraries in vitro.
An in vitro protein selection method, ribosome display, has been applied to comprehensively identify and map the immunologically relevant proteins of the human pathogen Staphylococcus aureus. A library built up from genomic fragments of the virulent S. aureus COL strain (methicillin-resistant S. aureus) allowed us to screen all possible encoded peptides for immunoreactivity. As selective agents, human sera exhibiting a high antibody titer and opsonic activity against S. aureus were used, since these antibodies indicate the in vivo expression and immunoreactivity of the corresponding proteins. Identified clones cluster in distinct regions of 75 genes, most of them classifiable as secreted or surface-localized proteins, including previously identified virulence factors. In addition, 14 putative novel short open reading frames were identified and their immunoreactivity and in vivo mRNA expression were confirmed, underscoring the annotation-independent, true genomic nature of our approach. Evidence is provided that a large fraction of the identified peptides cannot be expressed in an in vivo-based surface display system. Thus, in vitro protein selection, not biased by the context of living entities, allows screening of genomic expression libraries with a large number of different ligands simultaneously. It is a powerful approach for fingerprinting the repertoire of immune reactive proteins serving as target candidates for active and passive vaccination against pathogens.
Weichhart
Thomas
T
Intercell AG, 1030 Vienna, Austria.
Horky
Markus
M
Söllner
Johannes
J
Gangl
Susanne
S
Henics
Tamàs
T
Nagy
Eszter
E
Meinke
Andreas
A
von Gabain
Alexander
A
Fraser
Claire M
CM
Gill
Steve R
SR
Hafner
Martin
M
von Ahsen
Uwe
U
D000942:Antigens, Bacterial / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics* / Q000276:immunology*; D001483:Base Sequence; D004269:DNA, Bacterial / Q000235:genetics; D000939:Epitopes / Q000235:genetics; D015723:Gene Library; D016680:Genome, Bacterial; D006801:Humans; D066298:In Vitro Techniques; D016366:Open Reading Frames; D019151:Peptide Library; D010455:Peptides / Q000235:genetics / Q000276:immunology; D013209:Staphylococcal Vaccines / Q000235:genetics* / Q000302:isolation & purification; D013211:Staphylococcus aureus / Q000235:genetics* / Q000276:immunology* / Q000472:pathogenicity
D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D004269:DNA, Bacterial; D000939:Epitopes; D019151:Peptide Library; D010455:Peptides; D013209:Staphylococcal Vaccines