Properties of the N-terminal domains from Y receptors probed by NMR spectroscopy.
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.
Zou
Chao
C
Institute of Organic Chemistry, University of Zurich, Winterthurerstrasse 190, CH 8057 Zurich, Switzerland.
Kumaran
Sowmini
S
Walser
Reto
R
Zerbe
Oliver
O
D000595:Amino Acid Sequence; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D008969:Molecular Sequence Data; D009587:Nitrogen Isotopes; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D017476:Receptors, Neuropeptide Y / Q000737:chemistry*; D017386:Sequence Homology, Amino Acid
D009587:Nitrogen Isotopes; D017476:Receptors, Neuropeptide Y