Characterization of the recombinant Candida albicans β-1,2-mannosyltransferase that initiates the β-mannosylation of cell wall phosphopeptidomannan

• Published in: Biochemical journal Vol. 457; no. 2; p. 347
• Main Authors: Fabre, Emeline, Sfihi-Loualia, Ghenima, Pourcelot, Marilyne, Coddeville, Bernadette, Krzewinski, Frédéric, Bouckaert, Julie, Maes, Emmanuel, Hurtaux, Thomas, Dubois, Romaric, Fradin, Chantal, Mallet, Jean-Maurice, Poulain, Daniel, Delplace, Florence, Guerardel, Yann
• Format: Journal Article
• Language: English
• Published: England 15-01-2014
• Subjects: Candida albicans - enzymology; Candida albicans - genetics; Cell Wall - enzymology; Mannans - chemistry; Mannans - genetics; Mannans - metabolism; Mannose - chemistry; Mannose - genetics; Mannose - metabolism; Mannosyltransferases - chemistry; Mannosyltransferases - genetics; Mannosyltransferases - metabolism; Phosphopeptides - chemistry; Phosphopeptides - genetics; Phosphopeptides - metabolism; Recombinant Proteins - chemistry; Recombinant Proteins - genetics; Recombinant Proteins - metabolism
• ISSN: 1470-8728
• DOI: 10.1042/BJ20131012

The presence of β-mannosides in their cell walls confers specific features on the pathogenic yeasts Candida albicans and Candida glabrata compared with non-pathogenic yeasts. In the present study, we investigated the enzymatic properties of Bmt1 (β-mannosyltransferase 1), a member of the recently identified β-mannosyltransferase family, from C. albicans. A recombinant soluble enzyme lacking the N-terminal region was expressed as a secreted protein from the methylotrophic yeast Pichia pastoris. In parallel, functionalized natural oligosaccharides isolated from Saccharomyces cerevisiae and a C. albicans mutant strain, as well as synthetic α-oligomannosides, were prepared and used as potential acceptor substrates. Bmt1p preferentially utilizes substrates containing linear chains of α-1,2-linked mannotriose or mannotetraose. The recombinant enzyme consecuti-vely transfers two mannosyl units on to these acceptors, leading to the production of α-mannosidase-resistant oligomannosides. NMR experiments further confirmed the presence of a terminal βMan (β-1,2-linked mannose) unit in the first enzyme product. In the future, a better understanding of specific β-1,2-mannosyltransferase molecular requirements will help the design of new potential antifungal drugs.