Molecular mechanism of muscarinic acetylcholine receptor M3 interaction with Gq

Abstract

Muscarinic acetylcholine receptor M3 (M3) and its downstream effector Gq/11 are critical drug development targets due to their involvement in physiopathological processes. Although the structure of the M3-miniGq complex was recently published, the lack of information on the intracellular loop 3 (ICL3) of M3 and extensive modification of G alpha q impedes the elucidation of the molecular mechanism of M3-Gq coupling under more physiological condition. Here, we describe the molecular mechanism underlying the dynamic interactions between full-length wild-type M3 and Gq using hydrogen-deuterium exchange mass spectrometry and NanoLuc Binary Technology-based cell systems. We propose a detailed analysis of M3-Gq coupling through examination of previously well-defined binding interfaces and neglected regions. Our findings suggest potential binding interfaces between M3 and Gq in pre-assembled and functionally active complexes. Furthermore, M3 ICL3 negatively affected M3-Gq coupling, and the G alpha q AHD underwent unique conformational changes during M3-Gq coupling. Analysis of M3-Gq coupling utilized fulllength proteins and techniques like HDX-MS & NanoBiT assay, revealing potential binding interfaces in the preassembled and agonistactive complexes and critical regions for the complex formation and Gq activation.