Structural and functional characterization of allatostatin receptor type-C of Thaumetopoea pityocampa revealed the importance of Q271(6.55) residue in G protein-dependent activation pathway

Item Type:

Preprint

Title:

Creators Name:

Shahraki, A. and Isbilir, A. and Dogan, B. and Lohse, M.J. and Durdagi, S. and Birgul-Iyison, N.

Abstract:

Insect neuropeptide receptors are among the potential targets for designing next-generation pesticides. Activation of allatostatin receptor type C (AstR-C), a G Protein-coupled receptor (GPCR), upon stimulation with its endogenous ligand, allatostatin C (AST-C), leads to the inhibition of juvenile hormone (JH) secretion that consequently regulates physiology of insects. Here we conducted in silico and in vitro approaches to characterize the structure and function of AstR-C of Thaumetopoea pityocampa (T.pit), a well-known pest in Mediterranean countries. The sequence of AstR-C and AST-C were derived from whole genome sequencing (WGS) data. Resonance energy transfer (RET) methods were used to investigate the downstream effectors of the receptor and the temporal kinetics of G protein activation. Three-dimensional (3D) structure of AstR-C constructed via homology modeling methods was subjected to molecular dynamics (MD) simulations and docking studies to identify the orthosteric pocket. Our results showed that T.pit AstR-C couples to Gαi/o subtype of G proteins at sub-nanomolar ranges of the the ligand with the G protein recruitment and activation kinetics of ~4 and 6 seconds, respectively, when 1 nM AST-C is administered. At the increasing concentration of native ligand, βarrestin was shown to be recruited at nanomolar ranges the ligand. Docking and MD simulation studies revealed the importance of extracellular loop 2 (ECL2) in T.pit AstRC/AST-C interaction, and combination of in silico and in vitro methods supported the accuracy of the built model and the predicted orthosteric pocket. Q271(6.55) (Ballesteros-Weinstein generic numbering) was found to have a substantial role in G protein dependent activation of AstR-C possibly via contributing to the flexibility of the structure.

Keywords:

GPCR, Insect Neuropeptide Receptor, RET-Based Assays, MD Simulation, Molecular Docking Studies, Thaumetopoea pityocampa

Source:

bioRxiv

Publisher:

Cold Spring Harbor Laboratory Press

Article Number:

2020.07.01.174037

Date:

2 July 2020

Official Publication:

https://doi.org/10.1101/2020.07.01.174037

Related to:

URL	URL Type
https://edoc.mdc-berlin.de/19904/	Final version

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