Analysis of higher-order oligomers of melatonin receptors and their complexities

Abstract

The functions of melatonin are predominately mediated through two subtypes of human melatonin receptors, classified as melatonin type 1 (MT₁) and type 2 (MT₂), which utilize heterotrimeric G_i proteins for signaling. Although melatonin receptors can function as monomers, emerging biochemical and biophysical evidence has revealed that they form non-obligatory heterodimers. Since signals arising from the two melatonin receptor subtypes are not identical, one lingering question is whether the G_i protein exhibits any binding preference for the MT₁ or MT₂ protomer of the heterodimer, as this may facilitate or promote biased signaling properties. Despite prolific expansion in the number of receptor crystal structures being resolved, there is relatively limited molecular information about the precise configuration of dimeric receptors in complex with G proteins. Computational approaches provide a means to explore and visualize possible assembly of the heterodimeric form of melatonin receptors in complex with G_i protein, which is otherwise difficult to attain through contemporary biochemical or biophysical experimentation. Here, using RosettaDock and RosettaMPdock, molecular and structural insights on the signaling complex were gained by docking various combinations of MT₁, MT₂, and G_i protein against each other. Based on G_ibb’s free energy and dissociation constant values, the G_i protein was predicted to preferentially associate with the MT₁ protomer of the heterodimeric complex. Preferential signaling via the MT₁ protomer of the MT₁/MT₂ heterodimer was subsequently corroborated by cell-based functional assays where MT₁-specific stimulation of cAMP formation was demonstrated in HEK293 cells co-expressing the two receptors. Our results support the notion that stable heterodimeric receptor/G protein complexes can be formed prior to agonist binding, and only one G protein may be preferentially bound to one of the protomers in the melatonin receptor heterodimer, which in turn may predetermine the functional outcome upon receptor activation

Date of Award	2023
Original language	English
Awarding Institution	The Hong Kong University of Science and Technology
Supervisor	Yung Hou WONG (Supervisor)