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Glycosylation of the receptor guanylate cyclase C: role in ligand binding and catalytic activity

Ghanekar, Yashoda and Chandrashaker, Akhila and Tatu, Utpal and Viswewariah, Sandhya S (2004) Glycosylation of the receptor guanylate cyclase C: role in ligand binding and catalytic activity. In: Biochemical Journal, 139 (1). pp. 653-663.

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Official URL: http://www.ncbi.nlm.nih.gov/pmc/articles/PMC122412...

Abstract

GC-C (guanylate cyclase C) is the receptor for heat-stable enterotoxins, guanylin and uroguanylin peptides. Ligand binding to the extracellular domain of GC-C activates the guanylate cyclase domain leading to accumulation of cGMP. GC-C is expressed as differentially glycosylated forms in HEK-293 cells (human embryonic kidney-293 cells). In the present study, we show that the 145 kDa form of GC-C contains sialic acid and galactose residues and is present on the PM (plasma membrane) of cells, whereas the 130 kDa form is a high mannose form that is resident in the endoplasmic reticulum and serves as the precursor for the PM-associated form. Ligand-binding affinities of the differentially glycosylated forms are similar, indicating that glycosylation of GC-C does not play a role in direct ligand interaction. However, ligand-stimulated guanylate cyclase activity was observed only for the fully mature form of the receptor present on the PM, suggesting that glycosylation had a role to play in imparting a conformation to the receptor that allows ligand stimulation. Treatment of cells at 20 degrees C led to intracellular accumulation of a mature glycosylated form of GC-C that now showed ligand-stimulated guanylate cyclase activity, indicating that localization of GC-C was not critical for its catalytic activity. To determine if complex glycosylation was required for ligand-stimulated activation of GC-C, the receptor was expressed in HEK-293 cells that were deficient in N -acetylglucosaminyltransferase 1. This minimally glycosylated form of the receptor was expressed on the cell surface and could bind a ligand with an affinity comparable with the 145 kDa form of the receptor. However, this form of the receptor was poorly activated by the ligand. Therefore our studies indicate a novel role for glycosidic modification of GC-C during its biosynthesis, in imparting subtle conformational changes in the receptor that allow for ligand-mediated activation and perhaps regulation of basal activity.

Item Type: Journal Article
Additional Information: This article copyright belongs to Biochemical Society.
Department/Centre: Division of Biological Sciences > Molecular Reproduction, Development & Genetics (formed by the merger of DBGL and CRBME)
Date Deposited: 28 Jun 2007
Last Modified: 17 Jan 2012 09:54
URI: http://eprints.iisc.ernet.in/id/eprint/11312

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