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Complications in cell-surface labelling by biotinylation of Candida albicans due to avidin conjugate binding to cell-wall proteins

Departments of Pathology¹ and Microbiology², University of Virginia Health System, Charlottesville, VA 22908, USA

Author for correspondence: James Masuoka. Tel: +1 434 243 3744. Fax: +1 434 924 9312. e-mail: jm2n{at}virginia.edu

Initial contact between the opportunistic fungal pathogen Candida albicans and host tissue occurs at the cell surface. Biotin derivatives have been used to label the cell-surface proteins of yeasts, with labelled proteins subsequently detected by avidin–reporter conjugates. Previous work has indicated that avidin can bind to C. albicans proteins in the absence of biotin, suggesting a possible host-cell-recognition mechanism by fungal cell-surface proteins. To investigate this mechanism, Western blots of proteins extracted from biotinylated and mock-treated cells were probed with avidin or modified-avidin reagents. Each avidin reagent bound to cell-wall proteins extracted from non-biotinylated cells. Binding did not appear to be due to the lectin-like activity of the cell-wall proteins of C. albicans or to the presence of biotin in the sample itself. Binding was inhibited by added biotin, by the chaotrope KSCN and by NaCl in a concentration-dependent manner, although inhibition varied among the avidin conjugates tested. Thus, the non-specific binding of avidin to the cell-wall proteins of C. albicans appears to involve hydrophobic and electrostatic interactions, depending on the particular avidin species. These observations demonstrate potential pitfalls in the use of avidin–biotin complexes to identify cell-surface molecules and could provide insights into protein–protein interactions at the C. albicans cell wall.

Keywords: yeast, adhesion

Abbreviations: HRP, horseradish peroxidase; LYT, supernatant produced following treatment with lyticase; SDS extract, supernatant produced following SDS treatment of disrupted cell pellet; SUP, supernatant produced following cell disruption by glass beads

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