Solubilization, Purification, and Ligand Binding Characterization of G Protein-coupled Receptor SMO in Native Membrane Bilayer Using Styrene Maleic Acid Copolymer

Overview

Journal PeerJ

Specialties Biology
Environmental Health
General Medicine

Date 2022 May 9

PMID 35529497

Authors

Lina Zhu

Hongxin Zhao

Yizhuo Wang

Chuandi Yu

Juanjuan Liu

Ling Li

Zehua Li

Jin Zhang

Han Dai

Junfeng Wang

Lei Zhu

Affiliations

Soon will be listed here.

Abstract

Smoothened (SMO) protein is a member of the G protein-coupled receptor (GPCR) family that is involved in the Hedgehog (Hh) signaling pathway. It is a putative target for treating various cancers, including medulloblastoma and basal cell carcinoma (BCC). Characterizing membrane proteins such as SMO in their native state is highly beneficial for the development of effective pharmaceutical drugs, as their structures and functions are retained to the highest extent in this state. Therefore, although SMO protein is conventionally solubilized in detergent micelles, incorporating the protein in a lipid-based membrane mimic is still required. In this study, we used styrene maleic acid (SMA) copolymer that directly extracted membrane protein and surrounding lipids as well as formed the so-called polymer nanodiscs, to solubilize and purify the SMO transmembrane domain encapsulated by SMA-nanodiscs. The obtained SMA-nanodiscs showed high homogeneity and maintained the physiological activity of SMO protein, thereby enabling the measurement of the dissociation constant (K) for SMO ligands SMO-ligands Shh Signaling Antagonist V (SANT-1) and Smoothened Agonist (SAG) using ligand-based solution nuclear magnetic resonance spectroscopy. This work paves the way for investigating the structure, function, and drug development of SMO proteins in a native-like lipid environment.

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