Vesicular-integral membrane protein, VIP36, recognizes high-mannose type glycans containing α1→2 mannosyl residues in MDCK cells

Abstract

The 36 kDa vesicular-integral membrane protein, VIP36, has been originally isolated from MDCK cells as a component of glycolipid-enriched detergent-insoluble complexes containing apical marker proteins, and its luminal domain shows homology to leguminous plant lectins and ERGIC-53. As the first step to identify the functional role of VIP36, the carbohydrate binding specificity of VIP36 was investigated using a fusion protein of glutathione-S-transferase and luminal domain of VIP36 (Vip36). It was found that VIP36 recognizes high-mannose type glycans containing α1→2 Man residues and α-amino substituted asparagine. The binding of Vip36 to high-mannose type glycans was independent of Ca2+ and the optimal condition was pH 6.0 at 37°C. The concentration at which half inhibition of the binding by Man7-9·GlcNAc2·NAc·Asn occurred was 1.0 x 10-9 M. The association constant between Man7-9·GlcNAc2 in porcine thyroglobulin and immobilized Vip36 was 2.1 x 108 M-1 as determined by means of a biosensor based on surface plasmon resonance. These results indicate that VIP36 functions as an intracellular lectin recognizing glycoproteins which possess high-mannose type glycans, (Manα1→2)2-4·Man5·GlcNAc2.