Duggisetty, S. C.; Kumar, G.; Baratam, K.; Srivastava, A.

Sorting Nexins (SNXs) are a large group of diverse cellular trafficking proteins that play essential roles in membrane remodeling and cargo sorting between organelles. SNX proteins comprise a banana-shaped BAR domain that acts as a curvature-inducing scaffold and a phosphoinositide lipids sensitive Phox-Homology Domain (PXD) that interacts with the membrane to ensure specific and efficient organelle binding. In concert with the larger retromer machinery, these proteins traffic and recycle cargo between the endosomal membrane, trans-Golgi network, and plasma membrane. Interestingly, the SNX1-SNX5 heterodimeric construct forms a part of the newly discovered pathway where cargo sorting and membrane remodeling can take place in a retromer-independent fashion. In this work, we use molecular dynamics and continuum mechanics simulations to understand the features of SNX1-SNX5 heterodimer, especially the molecular determinants at PXDs, which impart organelle membrane specificity and retromer-independent cargo recognition ability to these proteins. Our all-atom molecular dynamics simulations with isolated PXDs and full-length SNX1-SNX5 on bilayers show that SNX1-PXD has robust membrane-binding features that are largely insensitive to single or double mutation of the basic residues on its surface. Comparing the simulation-based binding poses against the recently solved cryo-EM structures of tubular membrane-bound SNX1 homodimer and SNX1-SNX5 heterodimer also provided interesting insights into the association profile of isolated PXDs when they have the freedom to explore different membrane-binding poses. Our protein-protein simulations of SNX5-PXD with the tail region of the CI-MPR transmembrane cargo protein using metadynamics simulations reveal aromatic residue-rich {pi} - {pi} interactions between the two proteins, and a favorable and kinetically accessible binding free energy profile for SNX5. To model the emergent behavior of cargo sequestration and endosomal tube formation by SNX1-SNX5 heterodimer, we also performed Dynamically Triangulated Surface (DTS) based mesoscopic simulations by developing an augmented Helfrich-like continuum-mechanics Hamiltonian to incorporate transmembrane proteins in DTS models.