Biophysical Society Thematic Meeting | Trieste 2024

Emerging Theoretical Approaches to Complement Single-Particle Cryo-EM

Poster Abstracts

9-POS Board 9 STRUCTURAL INSIGHTS AND COMPUTATIONAL DESIGN OF LIGAND MEDIATED FUNCTION RECOVERY IN D408Y AND I423T MISSENSE MUTANTS OF THE HUMAN FOLLICLE-STIMULATING HORMONE RECEPTOR Eduardo Jardón-Valadez 1 ; Teresa Zariñan 2,3 ; Selvaraj Nataraja 4 ; Henry N Yu 5 ; Alfredo Ulloa Aguirre 2,3 ; 1 Metropolitan Autonomous University, Earth Resources Department, Lerma, Mexico 2 National University of Mexico, CIUDAD DE MEXICO, Mexico 3 Instituto Nacional de Ciencias Médicas y Nutrición SZ, Red de Apoyo a la Investigación, Ciudad de Mexico, Mexico 4 TocopheRx, Burlington, MA, USA 5 CanWell Pharma Inc, Woburn, MA, USA Follicle-stimulating hormone receptor (FSHR) plays an essential role in reproductive function. Mutations in the structure of this GPCR may result in impaired receptor expression and function due to misfolding and intracellular retention of the defective receptor, leading to reproductive disorders. The present study took advantage of the recent cryo-electron microscopy (Cryo-EM) structure of the FSHR, which was further refined applying molecular dynamics (MD) simulations to elucidate the structural dynamics and signal transduction mechanisms of the wild type receptor. Using these structures, we then applied computational biophysics approaches to test whether a novel allosteric agonist may restore expression and function of mutant FSHR variants. Our refined MD simulations revealed key conformational changes and interactions within the transmembrane (TM) regions of FSHR, highlighting potential allosteric binding sites critical for receptor activation. The small molecule (CAN1405; CanWell Pharma Inc., Woburn, MA, USA) showed promising binding affinity and efficacy for rescuing expression and function of FSHRs with clinically relevant missense mutations. Trafficking defective mutations in the ectodomain of the FSHR failed to rescue expression of the misfolded receptor in response to the allosteric agonist, whereas those mutations in the TM domain (mainly at TM2 and the extracellular loop 2) favored trafficking of the misfolded FSHR to the cell surface plasma membrane in response to the allosteric agonist. Biochemical studies showed that the rescued FSHRs responded to the orthosteric agonist in terms of cAMP production and phosphorylation of MAPK/ERK1/2. These findings provide a comprehensive structural and dynamic frame work for understanding FSHR signal transduction and offer a promising therapeutic strategy for treating mutation-induced receptor dysfunction. This study underscores the synergistic potential of Cryo-EM and computational biophysics in drug discovery and precision medicine, offering novel insights and new therapeutic avenues for reproductive health disorders. [Study supported by grant IN208323 from the PAPIIT-UNAM, Mexico (to A.U.-A.)].

41