Relative Abundance of Oligosaccharides in Candida Species as Determined by Fluorophore-Assisted Carbohydrate Electrophoresis

Abstract

ABSTRACT Fluorophore-assisted carbohydrate electrophoresis (FACE) is a straightforward, sensitive method for determining the presence and relative abundance of individual oligomannosyl residues in Candida mannoprotein, the major antigenic determinant located on the outer surface of the yeast cell wall. The single terminal aldehydes of oligomannosyl residues released by hydrolysis were tagged with the charged fluorophore 8-aminonaphthalene-1,3,6-trisulfonate (ANTS) and separated with high resolution on the basis of size by polyacrylamide gel electrophoresis. ANTS fluorescence labeling was not biased by oligomannoside length; therefore, band fluorescence intensity was directly related to the relative abundance of individual oligomannoside moieties in heterogeneous samples. FACE analysis revealed the major oligomannosides released by acid hydrolysis and β-elimination of Fehling-precipitated mannan from Candida albicans , which were the same as those previously reported in studies based on mass and nuclear magnetic spectroscopic analysis. FACE was also amenable to the analysis of samples obtained by direct hydrolysis of whole yeast cells. Whole-cell acid hydrolysis and whole-cell β-elimination of two isolates each of C. albicans , C. glabrata , C. krusei , C. lusitaniae , C. parapsilosis , C. rugosa , C. stellatoidea , and C. tropicalis resulted in oligomannoside gel banding patterns that were species and strain specific for the 16 isolates surveyed. Whereas some bands were specific for an individual isolate or species, other bands were shared by two or three species in various groupings. Differences in the mannoprotein composition of C. albicans A9 and four spontaneous cell surface mutants were also detected. Mannan “fingerprints,” or banding pattern profiles, derived from the electrophoretic mobilities of individual bands relative to the migration of acid-hydrolyzed dextran (relative migration index) yielded profiles characteristic of individual isolates not revealed by standard assimilation and biochemical profiles. FACE represents an accessible, sensitive, and quantitative analytical tool enabling the characterization of yeast mannan complexity.