Cloning and analysis of a Candida albicans gene that affects cell surface hydrophobicity.
The opportunistic pathogenic yeast Candida albicans exhibits growth phase-dependent changes in cell surface hydrophobicity, which has been correlated with adhesion to host tissues. Cell wall proteins that might contribute to the cell surface hydrophobicity phenotype were released by limited glucanase digestion. These proteins were initially characterized by their rates of retention during hydrophobic interaction chromatography--high-performance liquid chromatography and used as immunogens for monoclonal antibody production. The present work describes the cloning and functional analysis of a C. albicans gene encoding a 38-kDa protein recognized by the monoclonal antibody 6C5-H4CA. The 6C5-H4CA antigen was resolved by two-dimensional electrophoresis, and a partial protein sequence was determined by mass spectrometry analysis of tryptic fragments. The obtained peptides were used to identify the gene sequence from the unannotated C. albicans DNA database. The antibody epitope was provisionally mapped by peptide display panning, and a peptide sequence matching the epitope was identified in the gene sequence. The gene sequence encodes a novel open reading frame (ORF) of unknown function that is highly similar to several other C. albicans ORFs and to a single Saccharomyces cerevisiae ORF. Knockout of the gene resulted in a decrease in measurable cell surface hydrophobicity and in adhesion of C. albicans to fibronectin. The results suggest that the 38-kDa protein is a hydrophobic surface protein that meditates binding to host target proteins.
Singleton
D R
DR
Department of Pathology, University of Virginia Health Sciences Center, Charlottesville, Virginia 22908, USA.
Masuoka
J
J
Hazen
K C
KC
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D000946:Antigens, Fungal / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D001483:Base Sequence; D015153:Blotting, Western; D002176:Candida albicans / Q000166:cytology / Q000235:genetics* / Q000378:metabolism; D002448:Cell Adhesion; D002473:Cell Wall / Q000378:metabolism; D003001:Cloning, Molecular; D015180:Electrophoresis, Gel, Two-Dimensional; D018604:Epitope Mapping; D005353:Fibronectins / Q000378:metabolism; D005656:Fungal Proteins; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D017422:Sequence Analysis, DNA; D020539:Sequence Analysis, Protein; D013499:Surface Properties
D000911:Antibodies, Monoclonal; D000946:Antigens, Fungal; D001426:Bacterial Proteins; C430064:CSH1 protein, Candida albicans; D005353:Fibronectins; D005656:Fungal Proteins