Identification and cloning of GCA1, a gene that encodes a cell surface glucoamylase from Candida albicans.
Adherence of yeast cells of Candida albicans to human oesophageal cells is greater when cells are grown in 500 mM D-galactose in comparison to D-glucose at the same concentration. Moreover, a 190 kDa mannoprotein (MP190) from a yeast cell wall preparation is highly expressed when cells are grown in the presence of galactose but less so in glucose. We now report on the identification of the MP190 and the isolation of its encoding gene. MP190 was purified, and three internal peptides were isolated and sequenced. Each of the three peptides showed significant homology (65-85%) with a glucoamylase (GAM1) from the yeast, Schwanniomyces occidentalis. In order to isolate the C. albicans homologue of GAM1 (GCA1), we probed a genomic library with a 0.9-kb internal fragment of the S. occidentalis GAM1 and isolated a 2.3-kb clone that corresponded to the 5' region of the gene. Polymerase chain reaction (PCR) amplification was used to isolate the remainder of the open reading frame. GCA1 encodes a 946 amino acid protein containing three putative hydrophobic, membrane-spanning domains and 15 potential N-glycosylation sites. Both Gca1p and GAM1 are novel to the family of glycosyl hydrolases. Northern analysis indicated that GCA1 is transcribed to a greater extent in galactose than in sucrose or glucose. Also, using reverse transcriptase (RT)-PCR, we observed expression of GCA1 in a rat model of oral candidiasis, indicating that Gca1p is expressed during disease development.
Sturtevant
J
J
Georgetown University Medical Center, Department of Microbiology and Immunology, Washington, DC 20007, USA. sturtevj@gusun.georgetown.edu
Dixon
F
F
Wadsworth
E
E
Latge
J P
JP
Zhao
X J
XJ
Calderone
R
R
D000595:Amino Acid Sequence; D000818:Animals; D002176:Candida albicans / Q000201:enzymology* / Q000235:genetics; D002180:Candidiasis, Oral / Q000382:microbiology*; D003001:Cloning, Molecular; D004195:Disease Models, Animal; D005800:Genes, Fungal; D005087:Glucan 1,4-alpha-Glucosidase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000520:alpha-Glucosidases / Q000235:genetics
D000520:alpha-Glucosidases; D005087:Glucan 1,4-alpha-Glucosidase