Structural Studies of the Neuronal Apoptotic Complex of the Prongf-P75NTR-Sortilin

Abstract

One of the prominent causes of neurodegenerative diseases is the dysregulation of the p75 neurotrophin receptor (p75NTR)-mediated signaling. Mature nerve growth factor (NGF) binds to p75NTR and the co-receptor tyrosine kinase a (TrkA) to maintain the neuronal proliferation, whereas the precursor of NGF (proNGF) interacts with p75NTR and sortilin, leading to neuronal apoptosis. The imbalance of the two signaling activities has strongly associated with Alzheimer's Disease and neurodegeneration. To elucidate the molecular mechanism of the proNGF-p75NTR-sortilin complex formation, we generated the extracellular domains of the target proteins and characterized the protein bindings in vitro. The extracellular domain complex will show the key structural motifs for protein-protein interactions, since the extracellular domains drive the complex formation. We used the Sf9 insect cell to perform heterologous overexpression, maintaining the simple post-glycosylation for membrane proteins. Previous study showed that sortilin forms a dimer in an acidic environment. However, negative-stain electron microscopy showed that the ternary complex of the proNGF-p75NTR-sortilin was formed in a dimeric assembly and was not sensitive to the environmental pH. This is likely that the binding interface may not be via the interaction between sortilins. To unravel the mechanistic view and identify the key residues in the binding interface, we are pursuing single-particle electron cryogenic microscopy (cryo-EM) to determine a high-resolution structure of the ternary complex. The result will provide a structural insight into the development of new medical interventions for treating neurodegenerative disorders.