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Properties of the N-terminal domains from Y receptors probed by NMR spectroscopy


Zou, C; Kumaran, S; Walser, R; Zerbe, O (2009). Properties of the N-terminal domains from Y receptors probed by NMR spectroscopy. Journal of Peptide Science, 15(3):184-191.

Abstract

Binding of neurohormones from the NPY family to their receptors, the so-called Y receptors, that belong to the superfamily 1b of G-protein coupled receptors might include transient binding to the N-terminal domains of the receptors. Accordingly, we have studied structural features of the N-terminal domains from the Y1, Y2, Y4, and Y5 receptor subtypes (N-Y1, N-Y2, N-Y4, N-Y5). We developed efficient strategies for their recombinant expression. N-Y4 and N-Y1 were expressed as insoluble fusions to enforce accumulation into inclusion bodies, whereas N-Y2 and N-Y5 were expressed as soluble fusion proteins. All N-terminal
domains are fully flexible in aqueous buffer. In the presence of phospholipid micelles some stretches within the polypeptides adopt helical conformations, but these are too unstable to be characterized in detail. Using chemical shift mapping techniques, interactions of NPY, peptide YY (PYY), and pancreatic polypeptide (PP), the three members of the neurohormone family that are the Y receptors’ natural ligands, with N-Y1, N-Y2, and N-Y5 revealed chemical shift changes in all cases, with the largest values being encountered for PP interacting with N-Y1 or N-Y5 both in the presence and in the absence of phospholipid micelles. The strength of the interactions, however, is generally weak, and the data also point to nonspecific contacts. Previously, in case of the interaction of N-Y4 with PP, the contacts were shown to be electrostatic in nature. This work indicates that association of the peptides with the N-terminal domains may generally be part of their binding trajectory.

Abstract

Binding of neurohormones from the NPY family to their receptors, the so-called Y receptors, that belong to the superfamily 1b of G-protein coupled receptors might include transient binding to the N-terminal domains of the receptors. Accordingly, we have studied structural features of the N-terminal domains from the Y1, Y2, Y4, and Y5 receptor subtypes (N-Y1, N-Y2, N-Y4, N-Y5). We developed efficient strategies for their recombinant expression. N-Y4 and N-Y1 were expressed as insoluble fusions to enforce accumulation into inclusion bodies, whereas N-Y2 and N-Y5 were expressed as soluble fusion proteins. All N-terminal
domains are fully flexible in aqueous buffer. In the presence of phospholipid micelles some stretches within the polypeptides adopt helical conformations, but these are too unstable to be characterized in detail. Using chemical shift mapping techniques, interactions of NPY, peptide YY (PYY), and pancreatic polypeptide (PP), the three members of the neurohormone family that are the Y receptors’ natural ligands, with N-Y1, N-Y2, and N-Y5 revealed chemical shift changes in all cases, with the largest values being encountered for PP interacting with N-Y1 or N-Y5 both in the presence and in the absence of phospholipid micelles. The strength of the interactions, however, is generally weak, and the data also point to nonspecific contacts. Previously, in case of the interaction of N-Y4 with PP, the contacts were shown to be electrostatic in nature. This work indicates that association of the peptides with the N-terminal domains may generally be part of their binding trajectory.

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:07 Faculty of Science > Department of Chemistry
Dewey Decimal Classification:540 Chemistry
Uncontrolled Keywords:GPCR, structural biology, NMR
Language:English
Date:January 2009
Deposited On:17 Mar 2009 07:19
Last Modified:05 Apr 2016 13:11
Publisher:Wiley-Blackwell
ISSN:1075-2617
Funders:Swiss National Science Foundation; Grant Number: 3100A0-11173, Forschungskredit of the University of Zurich
Publisher DOI:https://doi.org/10.1002/psc.1102

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