Abstract

The Receptor for Advanced Glycation End-products (RAGE) is a multi-ligand cell surface receptor implicated in various inflammatory and degenerative diseases. Its function is critically dependent on its single-pass transmembrane (TM) domain, which is essential for dimerization and signal transduction across the cell membrane. To facilitate high-resolution structural and dynamic studies, we aimed to express and purify the TM domain of human RAGE (RAGE-TM) in a form suitable for Nuclear Magnetic Resonance (NMR) spectroscopy. We successfully cloned the RAGE-TM sequence and optimized its overexpression in *Escherichia coli* using an established protocol for membrane proteins. The protein was extracted from the bacterial membrane using a detergent system and purified to homogeneity via a combination of affinity and size-exclusion chromatography. The purified RAGE-TM was reconstituted into bicelles, and preliminary 2D [1H, 15N]-HSQC NMR spectra confirmed the protein's folded state and stability, demonstrating its suitability for detailed structural-dynamic analysis. This work provides a crucial foundation for understanding the molecular mechanisms of RAGE activation and offers a target for structure-based drug design.