Structure of the Lectin Mannose 6-Phosphate Receptor Homology (MRH) Domain of Glucosidase II, an Enzyme That Regulates Glycoprotein Folding Quality Control in the Endoplasmic Reticulum

Linda J Olson,Ramiro Orsi,Solana G Alculumbre,Francis C Peterson,Ivan D Stigliano,Armando J Parodi,Cecilia D'Alessio,Nancy M Dahms

doi:10.1074/jbc.m113.450239

Abstract

Here we report for the first time the three-dimensional structure of a mannose 6-phosphate receptor homology (MRH) domain present in a protein with enzymatic activity, glucosidase II (GII). GII is involved in glycoprotein folding in the endoplasmic reticulum. GII removes the two innermost glucose residues from the Glc3Man9GlcNAc2 transferred to nascent proteins and the glucose added by UDP-Glc:glycoprotein glucosyltransferase. GII is composed of a catalytic GIIα subunit and a regulatory GIIβ subunit. GIIβ participates in the endoplasmic reticulum localization of GIIα and mediates in vivo enhancement of N-glycan trimming by GII through its C-terminal MRH domain. We determined the structure of a functional GIIβ MRH domain by NMR spectroscopy. It adopts a β-barrel fold similar to that of other MRH domains, but its binding pocket is the most shallow known to date as it accommodates a single mannose residue. In addition, we identified a conserved residue outside the binding pocket (Trp-409) present in GIIβ but not in other MRHs that influences GII glucose trimming activity.

Highlights

Glucosidase II is an endoplasmic reticulum enzyme involved in quality control of glycoprotein folding
Does optimal glucosidase II (GII) deglucosylation activity require the presence of the full complement of mannose units on nascent glycoproteins and the four conserved amino acids of the GII␤ subunit that are known to be essential for the ability of the mannose 6-phosphate receptor homology (MRH) domains of mannose 6-phosphate receptors (MPRs) to bind mannose 6-phosphate (Man6-P)-containing lysosomal enzymes [15, 16] (Fig. 2B)
Why does GII differ from most other glycosidases for which their maximal activity does not require the presence of a lectinlike domain? We have shown previously that a decrease in N-glycan mannose content sharply decreases in vivo GII activity without affecting the activity of UDP-Glc:glycoprotein glucosyltransferase (UGGT), thereby prolonging the half-life of monoglucosylated glycans produced by UGGT [15]

Summary

Background

Glucosidase II is an endoplasmic reticulum enzyme involved in quality control of glycoprotein folding. Structure of Glucosidase II ␤ MRH Domain proposed to serve as a negative regulator of ER-associated degradation [8] In contrast to these two proteins, GII is an early player in N-glycan processing as it removes the two inner glucoses from all transferred glycans (residues l and m; see Fig. 1B) irrespective of the folding status of the protein. Does optimal GII deglucosylation activity require the presence of the full complement of mannose units on nascent glycoproteins and the four conserved amino acids of the GII␤ subunit that are known to be essential for the ability of the MRH domains of MPRs to bind mannose 6-phosphate (Man6-P)-containing lysosomal enzymes [15, 16] (Fig. 2B). We identified a residue (Trp-409) important for GII activity and propose two models of how it could be influencing GII␤-mediated enhancement of GII activity

EXPERIMENTAL PROCEDURES

RESULTS AND DISCUSSION

Constraint violations