Influence of outer region mannosylphosphorylation on N-glycan formation by Candida albicans: Normal acid-stable N-glycan formation requires acid-labile mannosylphosphate addition

Abstract

The pathogenic yeast Candida albicans produces large N-glycans with outer regions containing only mannose residues. The outer region comprises a primary branch with multiple secondary and tertiary branches. Tertiary branches are linked to secondary branches by phosphodiester bridges. In the current model of outer chain elongation in the genetically related yeast Saccharomyces cerevisiae, synthesis of the branches occurs sequentially, primary to tertiary. Thus, disruption of mannosylphosphorylation, the initial step in tertiary branch formation, should not affect primary or secondary branch production. Compared to its wild-type parent, a C. albicans mutant defective in tertiary branch mannosylphosphorylation (mnn4Delta/mnn4Delta) made outer regions with reduced susceptibility to low acid acetolysis treatment, suggesting that the secondary or primary region had been modified. Higher acid acetolysis conditions were required to release the secondary branches from the primary branches. The released secondary branches constitute the subset of the wild-type secondary branches that lack a phosphate group. In contrast, the acid-stable region of both wild-type and mnn4Delta S. cerevisiae strains required high acid acetolysis conditions to release the secondary branches, despite having smaller and less complex secondary and tertiary branches. These results suggest that the complex and longer secondary and tertiary branches of C. albicans affect the conformation of the acid-stable region to render it more susceptible to acetolysis which implies secondary and tertiary branch formation in C. albicans are interdependent events and occur concurrently, rather than sequentially.

Structure and Function of Glycosylated Tandem Repeats from Candida albicans Als Adhesins

“…The Glycam Biomolecule Builder (http: //glycam.ccrc.uga.edu) (19) was used to add the disaccharide ␣-D-mannose-(1-2)-␣-D-mannose-1␣ to solvent-exposed Ser and Thr hydroxyls, as predicted by Rosetta, in the top-scoring model of each repeat. This disaccharide represents an "average" C. albicans or S. cerevisiae O-glycoside and conforms to the expected average stoichiometry of glycosylation of TRs synthesized by S. cerevisiae (12,15,35). The same sugars were similarly added to the exposed hydroxyl groups of the top-scoring structure for the 72-residue, two-domain structure Als5-R1R2 (Als5p repeat sequences 1 and 2).…”

“…The same sugars were similarly added to the exposed hydroxyl groups of the top-scoring structure for the 72-residue, two-domain structure Als5-R1R2 (Als5p repeat sequences 1 and 2). N-glycosylation sites were derivatized with GlcNAc 2 Man 9 yeast core glycan (15,19). The folded and glycosylated domain models were subjected to brief Molecular Dynamics simulations to minimize energy.…”

“…This mean ratio of 2:1 is consistent with known O-glycosylation structures in C. albicans and S. cerevisiae. In C. albicans, the O-linked oligosaccharides include ␣-mannose, ␣1,2-linked mannobiose, and mannotriose (12,15).…”

“…These sites were modeled with a yeast core glycan of GlcNac 2 Man 9 , a minimal sized N-glycan in C. albicans (15). The result was a polar position for these glycans on the more hydrophilic end of the prolate ellipsoidal domains (Fig.…”

Candida albicans Primes TLR Cytokine Responses through a Dectin-1/Raf-1–Mediated Pathway

“…In contrast, the C. albicans mnn4 mutant, defective in tertiary branch mannosylphosphorylation (Fig. 3A) (17,22), has no effect on the direct induction of cytokine production. C. albicans pmr1 null mutant has a defect in phosphomannosylation, as well as in O-and N-linked glycosylation (18) (Fig.…”