The GH130 family of mannoside phosphorylases contains glycoside hydrolases that target beta-1,2 mannosidic linkages in Candida mannan

Fiona Cuskin (1) , Arnaud Baslé (1) , Simon Ladeveze (2) , Alison Day (1) , Harry Gilbert (1) , Gideon Davies (3) , Gabrielle Veronese (2) , Elisabeth Lowe (1)

1 Institute for Cell and Molecular Biosciences, The Medical School Newcastle University
2 LISBP - Laboratoire d'Ingénierie des Systèmes Biologiques et des Procédés
3 York Structural Biology Laboratory, Department of Chemistry

The depolymerization of complex glycans is an important biological process that is of considerable interest to environmentally relevant industries. Bêta-mannose is a major component of plant structural polysaccharides and eukaryotic N-glycans. These linkages are primarily cleaved by glycoside hydrolases, although a family of glycoside phosphorylases, GH130, have also been shown to target Bêta-1,2 and Bêta-1,4 mannosidic linkages. In these phosphorylases bond cleavage was mediated by a single displacement reaction in which phosphate functions as the catalytic nucleophile. A cohort of GH130 enzymes, however, lack the conserved basic residues that bind the phosphate nucleophile, and it was proposed that these enzymes function as glycoside hydrolases. Here we show that two Bacteroides enzymes, BT3780 and BACOVA_03624, which lack the phosphate binding residues are indeed Bêta-mannosidases that hydrolyse Bêta-1,2-mannosidic linkages through an inverting mechanism. As the genes encoding these enzymes are located in genetic loci that orchestrate the depolymerisation of yeast α-mannans, it is likely that the two enzymes target the Bêta-1,2- mannose residues that cap the glycan produced by Candida albicans. The crystal structure of BT3780 in complex with mannose bound in the -1 and +1 subsites showed a pair of glutamates, Glu227 and Glu268 hydrogen bond to O1 of Bêta-mannose, and either of these residues may function as the catalytic base. The candidate catalytic acid and the other residues that interact with the active site mannose are conserved in both GH130 mannoside phosphorylases and Bêta- 1,2-mannosidases. Functional phylogeny identified a conserved lysine, Lys199 in BT3780, as a key specificity determinant for Bêta-1,2-mannosidic linkages.

Mots clés: X-ray crystallography; gut symbiont; yeast; cell wall; phosphorylase; glycoside hydrolase

Domaines: Sciences du Vivant [q-bio]

Fiona Cuskin, Arnaud Baslé, Simon Ladeveze, Alison Day, Harry Gilbert, et al.. The GH130 family of mannoside phosphorylases contains glycoside hydrolases that target beta-1,2 mannosidic linkages in Candida mannan. Journal of Biological Chemistry, 2015, 290 (41), ⟨10.1074/jbc.M115.681460⟩. ⟨hal-01184120v2⟩