Identification of a 58-kilodalton cell surface fibrinogen-binding mannoprotein from Candida albicans

Abstract

Treatment of both yeast (blastoconidia) and hyphal (blastoconidia with germ tubes) cells of Candida albicans with 1-mercaptoethanol (1ME) releases a complex array of cell wall-bound proteins and glycoproteins. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western immunoblotting with fibrinogen-anti-fibrinogen antibody allowed the identification of a 58-kDa mannoprotein (mp58) in both extracts which specifically interacts with human fibrinogen. Treatment of intact cells with low concentrations of 1-glucanase (Zymolyase 20T) for short periods or with ,ME abolished or significantly reduced binding of fibrinogen. A rabbit polyclonal antiserum was raised against the purified mp58 species released by 1ME from germinated blastoconidia (PAb anti-mp58). By Western blotting, the antiserum cross-reacted with the homologous 58-kDa fibrinogen-binding mannoprotein present in I3ME extracts from blastoconidia, and by indirect immunofluorescence, the antiserum labelled both yeast cells and hyphae, yet reactivity was found primarily on the cell surface of filamentous forms. Immunostaining of human infected tissue sections with PAb anti-mp58 showed that the mp58 species is also expressed in vivo; in this case, the species is in the forms of both yeast and hyphal elements similarly labelled by the antiserum. Purified immunoglobulin G fraction from the antiserum did not alter the binding of fibrinogen as determined by a modified enzyme-linked immunosorbent assay and Western blotting. The N-and 0-glycosidically linked carbohydrates represent 18 to 20%o and 3 to 4%, respectively, of the molecular mass of the mp58. 0-linked sugar residues may be involved in the interaction of the molecule with fibrinogen.

Inhibition of Candida albicans attachment to extracellular matrix by antibodies which recognize hydrophobic cell wall proteins

“…For example, C. albicans wall proteins of 37, 60, 62, 67, and 68 kDa have been observed to bind laminin [15,16]. These molecular masses are very similar to those of proteins reported to bind ¢brinogen [14], ¢bronectin [12], C3d [11,13], and iC3b [48]. Although there is some evidence that this multiplicity is due to protein glycosylation [48], it has not been demonstrated that these proteins are structurally related to each other.…”

Clinical strains of Candida albicans express the surface antigen glyceraldehyde 3-phosphate dehydrogenase in vitro and in infected tissues

“…C. albicans clinical isolates from blood of patients with invasive candidiasis were propagated as blastoconidia in liquid synthetic minimal medium at 28³C, harvested, and stored at 4³C for 72^96 h (starvation period); starved blastoconidia were inoculated in fresh Lee medium and incubated for 6 h at 28³C to obtain cultures of blastoconidia or at 37³C for the formation of blastoconidia bearing germ tubes as reported previously [30,31]. Most blastoconidia from all clinical strains used were able to form germ tubes under these experimental conditions.…”

“…Protein and glycoprotein components of the walls were obtained from intact cells by treatment with Lmercaptoethanol (LME) [31,32]. Molecular species in the LME extracts were separated by SDS-PAGE, transferred to polyvinylidene di£uoride membranes (Millipore) and immunodetected with PAb anti-CA-GAPDH [5,6], as described [5,30,31].…”

“…The LME extracts were analyzed by SDS-PAGE and Western blotting using the PAb anti-CA-GAPDH as a probe. Among the numerous protein species extracted by LME from the wall of intact cells [30,31], the PAb anti-CA-GAPDH detected a 33-kDa protein species in extracts from both blastoconidia ( Fig. 2A) and germinated blastoconidia (Fig.…”

Adherence ofCandida albicansto host cells

“…A number of plasma and extracellular matrix proteins have been demonstrated to bind to C. albicans. These include fibrinogen [45], vitronectin [46], laminin [47], and entactin [48]. Whether or not these receptors prove to be involved in the adherence process remains to be demonstrated.…”