THE GLYCOLYTIC ENZYME GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE OF CANDIDA-ALBICANS IS A SURFACE-ANTIGEN

A lambda gt11 cDNA library from Candida albicans ATCC 26555 Has screen ed by using pooled sera from two patients with systemic candidiasis an d five neutropenic patients with high levels of anti-C. albicans immun oglobulin M antibodies, Seven clones were isolated from 60,000 recombi nant phages. The most reactive one contained a 0.9-kb cDNA encoding a polypeptide immunoreactive only with sera from patients with systemic candidiasis, The whole gene was isolated from a genomic library by usi ng the cDNA as a probe. The nucleotide sequence of the coding region s howed homology (78 to 79%) to the Saccharomyces cerevisiae TDH1 to TDH 3 genes coding for glyceraldehyde-3-phosphate dehydrogenase (GAPDH), a nd their amino acid sequences showed 76% identity: thus, this gene has been named C. albicans TDH1. A rabbit polyclonal antiserum against th e purified cytosolic C. albicans GAPDH (polyclonal antibody [PAb] anti -CA-GAPDH) was used to identify the GAPDH in the beta-mercaptoethanol extracts containing cell wall moieties, Indirect immunofluorescence de monstrated the presence of GAPDH at the C. albicans cell surface, part icularly on the blastoconidia. Semiquantitative flow cytometry analysi s showed the sensitivity of this GAPDH form to trypsin and its resista nce to be removed with 2 M NaCl or 2% sodium dodecyl sulfate, The decr ease in fluorescence in the presence of soluble GAPDH indicates the sp ecificity of the labelling, In addition, a dose-dependent GAPDH enzyma tic activity was detected in intact blastoconidia and germ tube cells. This activity was reduced by pretreatment of the cells with trypsin, formaldehyde, and PAb anti-CA-GAPDH. These observations indicate that an immunogenic, enzymatically active cell wall-associated form of the glycolytic tic enzyme GAPDH is found at the cell surface face of C. al bicans cells.