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Re-evaluation of receptor–ligand interactions of the human neuropeptide Y receptor Y1: a site-directed mutagenesis study
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
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2006 (English)In: Biochemical Journal, ISSN 0264-6021, Vol. 1, no 393, 161-169 p.Article in journal (Refereed) Published
Abstract [en]

Interactions of the human NPY (neuropeptide Y) receptor Y1 with the two endogenous agonists NPY and peptide YY and two non-peptide antagonists were investigated using site-directed mutagenesis at 17 positions. The present study was triggered by contradictions among previously published reports and conclusions that seemed inconsistent with sequence comparisons across species and receptor subtypes. Our results show that Asp287, at the border between TM (transmembrane) region 6 and EL3 (extracellular loop 3) influences peptide binding, while two aspartic residues in EL2 do not, in agreement with some previous studies but in disagreement with others. A hydrophobic pocket of the Y1 receptor consisting of Tyr100 (TM2), Phe286 (TM6) and His298 (EL3) has been proposed to interact with the amidated C-terminus of NPY, a theory that is unsupported by sequence comparisons between Y1, Y2 and Y5. Nevertheless, our results confirm that these amino acid residues are critical for peptide binding, but probably interact with NPY differently than proposed previously. Studies with the Y1-selective antagonist SR120819A identified a new site of interaction at Asn116 in TM3. Position Phe173 in TM4 is also important for binding of this antagonist. In contrast with previous reports, we found that Phe173 is not crucial for the binding of BIBP3226, another selective Y1 receptor antagonist. Also, we found that position Thr212 (TM5) is important for binding of both antagonists. Our mutagenesis results and our three-dimensional model of the receptor based on the high-resolution structure of bovine rhodopsin suggest new interactions for agonist as well as antagonist binding to the Y1 receptor.

Place, publisher, year, edition, pages
2006. Vol. 1, no 393, 161-169 p.
Keyword [en]
BIBP3226, G-protein-coupled receptor (GPCR), mutagenesis, neuropeptide Y (NPY), receptor Y1, three-dimensional model
National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:uu:diva-97977DOI: 10.1042/BJ20050708PubMedID: 16097949OAI: oai:DiVA.org:uu-97977DiVA: diva2:173124
Available from: 2009-01-30 Created: 2009-01-30 Last updated: 2009-11-04Bibliographically approved
In thesis
1. Functional Studies of the Neuropeptide Y System: Receptor-Ligand Interaction and Regulation of Food Intake
Open this publication in new window or tab >>Functional Studies of the Neuropeptide Y System: Receptor-Ligand Interaction and Regulation of Food Intake
2009 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The members of the mammalian neuropeptide Y family, i.e. the peptides neuropeptide Y (NPY), peptide YY (PYY) and pancreatic polypeptide (PP), are all involved in regulation of food intake. In human and most other mammals they act via receptors Y1, Y2, Y4 and Y5. NPY is released in the hypothalamus and is one of the strongest appetite-stimulating neurotransmitters whereas PP and PYY are secreted from gut endocrine cells after meals and function as appetite-reducing hormones. This thesis describes studies of the NPY system at both the molecular and the physiological level. The first part describes two investigations of receptor-ligand interactions with the human Y1 and Y2 receptors. The results clarify the importance of several amino-acid residues of the human Y1 receptor. Three amino acids previously suggested by others to form a binding pocket for the carboxy-terminus of the peptide were confirmed to be crucial for interaction with peptide ligands. However, they were found to be too distantly located from each other to be able to form a binding pocket. Further investigation of the three corresponding positions in the human Y2 receptor showed that only one of the positions was important for interaction with full-length peptides. The results indicate overlapping but, surprisingly, non-identical binding of the different peptides to human Y1 and Y2 receptors, despite the fact that the two receptors share a common ancestor. The second part of the thesis describes an investigation of the effect of PP on food intake in six beagle dogs and a test for personality characteristics in dogs (TFPC). Treatment with physiological doses of PP decreased both the appetitive and the consummatory drive but had no effect on the amount food consumed. The TFPC protocol was used to map individual behavioral differences in a population of sixteen beagle dogs. The test, which included several situations that may appear in an experimental study, revealed considerable inter-individual differences in behavioral responses despite the fact that the dogs were born and housed in the same animal facility in constant controlled conditions. These results demonstrate that PP can influence food intake in distantly related mammals and emphasize the importance of considering differences in personality in experimental animals.

Place, publisher, year, edition, pages
Uppsala: Universitetsbiblioteket, 2009. 67 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 415
Keyword
neuropeptide Y, G-protein coupled receptor (GPCR), site-directed mutagenesis, pancreatic polypeptide, food intake, dog
National Category
Pharmacology and Toxicology
Identifiers
urn:nbn:se:uu:diva-9533 (URN)978-91-554-7393-8 (ISBN)
Public defence
2009-02-20, B21, BMC, Husargatan 3, Uppsala, 13:15
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Supervisors
Available from: 2009-01-30 Created: 2009-01-30 Last updated: 2013-01-07Bibliographically approved

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