Contributions of Glycosaminoglycans to Membrane Adsorption and Conformational Conversion of Peptide Hormones

Abstract

The membrane catalysis hypothesis advocates that a peptide ligand initially interacts with the plasma membrane and this interaction increases the local concentration at the vicinity of its cognate receptor. Besides, upon membrane adsorption, the peptide undergoes a conformational transition that favors receptor binding. In this mechanism, the lipidic components are the core of the membrane catalysis hypothesis. However, the membrane environment is complex and is constituted of a variety of macromolecules, including glycosaminoglycans (GAGs). Although these long polysulfated heteropolysaccharide chains have be shown to participate in the recruitment of chemokines, nothing is known about the contribution of GAGs in membrane adsorption of cationic peptidic hormones. In this context, we probed the contributions of GAGs in the membrane adsorption of secretin. This 27-residue peptide is secreted by the S cells of duodenum and known for regulating water homeostasis. To investigate the role of GAGs in the cell surface adsorption of secretin, we used CHO pgs-A-745 cells, which lack the first enzyme for GAGs biosynthesis, and their wild type counterpart, CHO K1. By confocal microscopy and flow cytometry, we observed that CHO pgs A-745 cells absorbed less fluorescein-labelled peptides than CHO-K1 cells. Then, we characterized the interaction between GAGs and secretin by affinity chromatography, isothermal titration calorimetry and surface plasmon resonance. A strong interaction between the peptide and GAGs was observed with a Kd in the low μM. Interestingly, heparin stabilizes the helical conformation of secretin. By designing a secretin analog with a restricted conformational ensemble, we observed that the spatial distribution of basic residues constitutes a key element for high affinity binding. Overall, this study redefines the membrane catalysis hypothesis and suggests that cationic peptide hormones initially bind to GAGs allowing a possible key role in receptor binding.