Isolation and preliminary characterization of the 14- to 18-kilodalton Candida albicans antigen as a phospholipomannan containing beta-1,2-linked oligomannosides.

AUTOR(ES)

Trinel, P A

RESUMO

Western blot (immunoblot) analysis of Candida albicans germ tube extracts has demonstrated the probable presence of beta-1,2-linked oligomannosides acting as epitopes distributed over a 14- to 18-kDa antigen unreactive to concanavalin A. These conclusions about the existence of these non-mannan-associated oligomannoside species were reinforced in the present study by the demonstration of reactivity of factor serum 5 (Iatron Laboratories) with the same antigen. A monoclonal antibody which reacted in an enzyme immunoassay with beta-1,2-linked oligomannosides converted into neoglycolipids and in Western blotting with the 14- to 18-kDa antigen from yeast and germ tubes, through metaperiodate-sensitive epitopes, was used for further characterization of the molecule. Reducing agents and strong protease digestion, which have deleterious effects on C. albicans proteins and mannoproteins, affected neither the antigenicity nor the relative molecular weight of the molecule. Western blots performed after migration of protease-treated extracts in polyacrylamide gels without sodium dodecyl sulfate (SDS) showed that the 14- to 18-kDa antigen could be negatively charged, whereas metabolic radiolabeling demonstrated that these charges could originate, at least in part, from the presence of phosphorus within the molecule. Chloroform-methanol-water extraction of protease-resistant material led to purification of the 14- to 18-kDa antigen, as determined by SDS-polyacrylamide gel electrophoresis and Western blotting. Metabolic radiolabeling with mannose confirmed the presence of these sugar residues within the purified 14- to 18-kDa antigen (despite its nonreactivity to concanavalin A), whereas radiolabeling with palmitic acid demonstrated its lipopolysaccharidic nature. Together, these results led to the conclusion that the 14- to 18-kDa antigen is a phospholipomannan.

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