Sugar-binding activity of the MRH domain in the ER alpha-glucosidase II beta subunit is important for efficient glucose trimming.

Glucosidase II (GII) is a glycan-processing enzyme that trims two alpha1,3-linked glucose residues from N-glycan on newly synthesized glycoproteins. Trimming of the first alpha1,3-linked glucose from Glc(2)Man(9)GlcNAc(2) (G2M9) is important for a glycoprotein to interact with calnexin/calreticulin (CNX/CRT), and cleavage of the innermost glucose from Glc(1)Man(9)GlcNAc(2) (G1M9) sets glycoproteins free from the CNX/CRT cycle and allows them to proceed to the Golgi apparatus. GII is a heterodimeric complex consisting of a catalytic alpha subunit (GIIalpha) and a tightly associated beta subunit (GIIbeta) that contains a mannose 6-phosphate receptor homology (MRH) domain. A recent study has suggested a possible involvement of the MRH domain of GIIbeta (GIIbeta-MRH) in the glucose trimming process via its putative sugar-binding activity. However, it remains unknown whether GIIbeta-MRH possesses sugar-binding activity and, if so, what role this activity plays in the function of GII. Here, we demonstrate that human GIIbeta-MRH binds to high-mannose-type glycans. Frontal affinity chromatography revealed that GIIbeta-MRH binds most strongly to the glycans with the alpha1,2-linked mannobiose structure. GII with the mutant GIIbeta that lost the sugar-binding activity of GIIbeta-MRH hydrolyzes p-nitrophenyl-alpha-glucopyranoside, but the capacity to remove glucose residues from G1M9 and G2M9 is significantly decreased. Our results clearly demonstrate the capacity of the GIIbeta-MRH to bind high-mannose-type glycans and its importance in efficient glucose trimming of N-glycans.