Molecular Basis of Sugar Recognition by the Human L-type Lectins ERGIC-53, VIPL, and VIP36

Yukiko Kamiya,Daiki Kamiya,Kazuo Yamamoto,Beat Nyfeler,Hans-Peter Hauri,Koichi Kato

doi:10.1074/jbc.m709384200

Abstract

ERGIC-53, VIPL, and VIP36 are related type 1 membrane proteins of the mammalian early secretory pathway. They are classified as L-type lectins because of their luminal carbohydrate recognition domain, which exhibits homology to leguminous lectins. These L-type lectins have different intracellular distributions and dynamics in the endoplasmic reticulum-Golgi system of the secretory pathway and interact with N-glycans of glycoproteins in a Ca(2+)-dependent manner, suggesting a role in glycoprotein sorting and trafficking. To understand the function of these lectins, knowledge of their carbohydrate specificity is crucial but only available for VIP36 (Kamiya, Y., Yamaguchi, Y., Takahashi, N., Arata, Y., Kasai, K. I., Ihara, Y., Matsuo, I., Ito, Y., Yamamoto, K., and Kato, K. (2005) J. Biol. Chem. 280, 37178-37182). Here we provide a comprehensive and quantitative analysis of sugar recognition of the carbohydrate recognition domains of ERGIC-53 and VIPL in comparison with VIP36 using a pyridylaminated sugar library in conjunction with frontal affinity chromatography. Frontal affinity chromatography revealed selective interaction of VIPL and VIP36 with the deglucosylated trimannose in the D1 branch of high-mannose-type oligosaccharides but with different pH dependence. ERGIC-53 bound high-mannose-type oligosaccharides with low affinity and broad specificity, not discriminating between monoglucosylated and deglucosylated high-mannosetype oligosaccharides. Based on the sugar-binding properties in conjunction with known features of these proteins, we propose a model for the action of the three lectins in glycoprotein guidance and trafficking. Moreover, structure-based mutagenesis revealed that the sugar-binding properties of these L-type lectins can be switched by single amino acid substitutions.

Highlights

The frontal affinity chromatography (FAC) data were collected for ER-Golgi intermediate compartment (ERGIC)-53-carbohydrate recognition domain (CRD) and VIP36-CRD
The results show that the sugar-binding specificity pattern of the VIPL-CRD is strikingly similar to that of the VIP36-CRD
A key finding is that the pH dependence of sugar binding was considerably different for VIP36 and VIPL (Fig. 2)

Summary

Introduction

In the present study the recombinant CRDs were immobilized through their histidine tag for FAC analyses except for experiments to examine pH dependence of sugar binding of VIPL and VIP36 (and its mutant). Sugar-binding Properties—Fig. 1B shows typical elution profiles of a series of PA oligosaccharides after application to the immobilized VIPL-CRD column at pH 7.4.

Results

Conclusion