Exploring the distinct activation mechanisms of neuromedin B receptor through multiple replica molecular dynamics simulations and Markov state modeling.

Neuromedin B receptor (NMBR) is a prototypical class A G protein-coupled receptor (GPCR) that plays a crucial role in histamine-independent itch transmission. However, the mechanisms underlying NMBR's selectivity and activation remain poorly understood. Herein, we utilized multiple replica molecular dynamics simulations, Markov state model (MSM), and community network analysis to explore how NMBR interacts with its two orthosteric ligands namely NMB30 and GRP(14-27) and the distinct activation mechanisms of NMBR. Our findings revealed distinct binding modes for these two ligands, highlighting the significance of the C-terminal dumbbell structure of peptides in ligand positioning and interaction with the "toggle switch" residue W279^6.48. Moreover, MSM analysis of the NMBR-GRP(14-27) complex revealed that the active conformation of the receptor appeared for less than half of the simulation time, confirming GRP(14-27) as a weak agonist of NMBR. Additionally, community network analysis identified community 8 as a connector between the orthosteric pocket and the intracellular end of NMBR, suggesting that ligand binding patterns may further influence the extent of receptor activation through allosteric regulation. Collectively, our study not only paves the way for the development of antipruritic therapy, but also provides valuable insights for future studies on GPCR activation.