# Characterization of [3H]Propionylated Human Peptide YY-A New Probe for Neuropeptide Y Y2 Receptor Binding Studies

## TL;DR

This paper introduces a new tritium-labeled PYY derivative for studying Y2R receptor binding, which works well in sodium-containing buffers.

## Contribution

A new tritiated PYY derivative is developed for Y2R binding studies in sodium-containing buffers.

## Key findings

- The [3H]propionylated PYY derivative shows saturable binding to Y2R in both sodium-free and sodium-containing buffers.
- Ki values from competition experiments align with known Y2R binding affinities, validating the new radioligand.
- The radioligand is suitable for Y2R binding assays in physiological sodium concentrations.

## Abstract

The neuropeptide
Y (NPY) Y2 receptor (Y2R)
is a G-protein-coupled receptor that is involved in the regulation
of various physiological processes such as neurotransmitter release,
bone metabolism, and memory. Consequently, the Y2R represents
a potential drug target, e.g., for the treatment of epilepsy and mood
disorders. Until now, the determination of the Y2R binding
affinities of Y2R ligands has primarily been performed
using 125I-labeled derivatives of the endogenous Y2R agonists NPY and peptide YY (PYY). A tritium-labeled NPY
derivative has also been used; however, its suitability for binding
assays in sodium-containing buffer is doubtful. We synthesized a tritium-labeled
PYY derivative by [3H]propionylation at Lys4 ([3H]2). The radioligand was characterized
by saturation binding, association, and dissociation kinetics and
was applied in competition binding assays. Specific binding of [3H]2 at intact Chinese hamster ovary cells expressing
the hY2R was saturable in both sodium-free buffer (apparent Kd = 0.016–0.067 nM) and sodium-containing
buffer (175 mM Na+, apparent Kd = 0.16–0.18 nM). Competition binding experiments with Y2R reference ligands yielded Ki values, which are in good agreement with the reported Y2R binding affinities, showing that [3H]2 represents
a useful tritiated tool compound for the determination of Y2R binding affinities also in buffers containing sodium at physiological
concentrations.