title: Expression of transglutaminase substrate activity on Candida albicans germ tubes through a coiled, disulfide-bonded N-terminal domain of Hwp1 requires C-terminal glycosylphosphatidylinositol modification.
abstract: By serving as a microbial substrate for epithelial cell transglutaminase, Hwp1 (Hyphal wall protein 1) of Candida albicans participates in cross-links with proteins on the mammalian mucosa. Biophysical properties of the transglutaminase substrate domain were explored using a recombinant protein representative of the N-terminal domain of Hwp1 and were similar to other transglutaminase substrates, the small proline-rich proteins of cornified envelopes found in stratified squamous epithelia. Recombinant Hwp1 lacks alpha and beta structures by circular dichroism and likely exists as a disulfide-cross-linked coiled-coil. The transglutaminase substrate property prompted a unique approach for investigating the features of surface Hwp1 on germ tubes. A lysine analog, 5-(biotinamido)pentylamine, was cross-linked to germ tubes catalyzed by transglutaminase 2 prior to cell fractionation, immunoprecipitation, and detection with streptavidin conjugates. The majority of the transglutaminase-modifiable Hwp1 was covalently attached to the beta-glucan of hyphae by the C terminus of Hwp1 via a glycosylphosphatidylinositol remnant anchor. A putative precursor of cell wall forms of Hwp1 was identified in the cell extract and in the culture medium. Hwp1 was modified by relatively short N-linked glycans, and the molecular size of the protein was reduced by hypomannosylation when expressed in O-glycosylation mutant strains. Hwp1 combines features of mammalian transglutaminase substrate proteins with characteristics of fungal cell wall proteins to form an unconventional adhesin at the hyphal wall of C. albicans.
authors.affiliation: Departments of Molecular Virology, Immunology, and Medical Genetics, The Ohio State University, Columbus, Ohio 43201, USA.
authors.forename: Janet F
authors.initials: JF
authors.lastname: Staab
authors.forename: Yong-Sun
authors.initials: YS
authors.lastname: Bahn
authors.forename: Chia-Hui
authors.initials: CH
authors.lastname: Tai
authors.forename: Paul F
authors.initials: PF
authors.lastname: Cook
authors.forename: Paula
authors.initials: P
authors.lastname: Sundstrom
mesh_terms: D000588:Amines / Q000737:chemistry; D000818:Animals; D055592:Biophysical Phenomena; D001703:Biophysics; D001710:Biotin / Q000031:analogs & derivatives* / Q000737:chemistry; D015153:Blotting, Western; D002176:Candida albicans / Q000201:enzymology*; D002448:Cell Adhesion; D002473:Cell Wall / Q000737:chemistry / Q000378:metabolism; D002942:Circular Dichroism; D004220:Disulfides / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D004847:Epithelial Cells / Q000378:metabolism; D005656:Fungal Proteins / Q000737:chemistry* / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000737:chemistry; D006026:Glycoside Hydrolases / Q000378:metabolism; D006031:Glycosylation; D017261:Glycosylphosphatidylinositols / Q000737:chemistry*; D006168:Guinea Pigs; D008099:Liver / Q000378:metabolism; D008239:Lysine / Q000737:chemistry; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000378:metabolism; D009154:Mutation; D016366:Open Reading Frames; D010716:Phosphatidylinositols / Q000737:chemistry; D010843:Pichia / Q000378:metabolism; D011233:Precipitin Tests; D011392:Proline / Q000737:chemistry; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry; D013050:Spectrometry, Fluorescence; D019809:Streptavidin / Q000737:chemistry; D013347:Subcellular Fractions; D013438:Sulfhydryl Compounds / Q000737:chemistry; D011503:Transglutaminases / Q000737:chemistry* / Q000378:metabolism
chemical_list: D000588:Amines; D004220:Disulfides; D005656:Fungal Proteins; D017261:Glycosylphosphatidylinositols; C118102:HWP1 protein, Candida albicans; D008562:Membrane Glycoproteins; D010716:Phosphatidylinositols; D011994:Recombinant Proteins; D013438:Sulfhydryl Compounds; C076022:5-(biotinamido)pentylamine; D001710:Biotin; D019809:Streptavidin; D011392:Proline; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D011503:Transglutaminases; D006026:Glycoside Hydrolases; D019204:GTP-Binding Proteins; D008239:Lysine