Identification of Continuous B-Cell Epitopes on the Protein Moiety of the 58-Kilodalton Cell Wall Mannoprotein ofCandida albicansBelonging to a Family of Immunodominant Fungal Antigens

Abstract

The 58-kiloDalton mannoprotein (mp58) on the surface of Candida albicans is highly immunogenic, is expressed by all C. albicans isolates tested, and elicits strong antibody responses during candidiasis. It belongs to a family of immunodominant fungal antigens with representatives also in different species of Aspergillus. The amino acid sequence of the protein portion of mp58 as deduced from the DNA sequence of its encoding gene (FBP1/PRA1) was used to synthesize a complete set of overlapping dodecapeptides (overlap, 7; offset, 5) covalently attached to the surface of derivatized polyethylene pins. The pin-coupled peptides were used in a modified enzyme-linked immunosorbent assay (ELISA) to identify continuous epitopes recognized by a number of antiserum preparations containing anti-mp58 antibodies. This comprehensive epitope-scanning study revealed the presence of multiple immunoreactive continuous B-cell epitopes within the protein sequence. Regions of increased reactivity included both the amino and carboxy termini of the mature protein (encompassing amino acid residues 16 to 50 and 286 to 299, respectively) and four internal regions spanning amino acids at positions 66 to 92, 121 to 142, 148 to 192, and 211 to 232. Further delineation of epitopic regions and identification of the boundaries of the antigenic sites was performed upon ELISA testing with a second Pepset consisting of completely overlapping 8-mer peptides spanning these reactive regions in the protein moiety of mp58. The highly reactive epitopic region at the C terminus of the protein was further evaluated using both window net and replacement net analyses. A synthetic peptide corresponding to the last 10 amino acid residues at the C terminus of the protein was immunogenic when injected into mice after being coupled to a carrier protein. Moreover, antibodies in the resulting sera specifically recognized the homologus mp58 in ELISAs and immunoblot assays. Delineation of the antibody responses to mp58 could provide the basis for the development of novel immunity-based prophylactic, therapeutic, and diagnostic techniques for the management of candidiasis.Candida albicans is both a commensal and an opportunistic pathogen of humans. Depending on the underlying host defect, this microorganism is able to cause a variety of infections that range from mucosal to life threatening disseminated candidiasis. C. albicans pathogenicity also depends on a complex array of microorganism-related virulence factors (reviewed in reference 15

Mapping of B-Cell Epitopes on a Novel 11.5-KilodaltonFasciola hepatica-Schistosoma mansoniCross-Reactive Antigen Belonging to a Member of theF. hepaticaSaposin-Like Protein Family

“…Previous studies relating to the epitope mapping of Mycoplasma bovis surface lipoproteins have shown B-cell epitopes to be around three to seven amino acid residues long (22). However, other studies have shown reactive B-cell epitopes relating to fungal, bacterial, or parasitical antigens to be as long as 10 or even 15 residues (15,27,28,29,30), similar to the size of the 10-mer SKQPTIDIDL, ADQTVEEHIG, or QQEVDYIIDH described here.…”

Analysis ofPRA1and Its Relationship toCandida albicans- Macrophage Interactions

“…The modulation of confirmed or putative cell surface/membrane/ secreted protein-encoding genes, such as PRA1, DDR48, CSH1, ECM4, HGT12, HGT18, CIP1, INO1, etc., can offer new insight into the molecular mechanisms involved in macrophage phagocytosis and/or Candida escape. One of the most highly upregulated genes identified in the pathogen upon engulfment by the macrophage was PRA1, a gene encoding a cell surface product that has been described as a fibrinogen binding protein (78). In vitro, the PRA1 (pH-regulated antigen 1) gene is activated, along with PHR1 (pH responsive) and its regulator RIM101 (regulator of IME2), when Candida cells are grown at an alkaline pH (15).…”

“…Pra1p is a cell wall-associated protein found in both the yeast and hyphal forms of C. albicans: Pra1p becomes highly glycosylated in hyphal-form cells (12) and elicits a strong immune response in infected patients (77). Pra1p was also identified as mp58 (58-kDa mannoprotein) or Fbp1 (fibrinogen binding protein) (45,78). An attractive model for a fibrinogen-binding role of Pra1p would be to link Candida cells to the Mac-1 receptor through fibrinogen (21,22,74), so the expression of this protein during phagocytosis could facilitate Candida adsorption to and escape from macrophages.…”

Saccharomyces uvarum mannoproteins stimulate a humoral immune response in mice

“…Another MP of 58 kDa with immunomodulatory activity has been identified in C. albicans. It belongs to a family of fungal immunogenic MPs and stimulates a high production of specific antibodies (Viudes et al 2001). The main mechanism by which the MP produces the modulation of the immune response is related to mannose receptors present on dendritic cells (Mansour et al 2006;Fernández et al 2005).…”