Structural Basis of Complex Formation Between Mitochondrial Anion Channel VDAC1 and Hexokinase-II

Abstract

ABSTRACTComplex formation between hexokinase-II (HKII) and the mitochondrial channel VDAC1 plays a crucial role in regulating cell growth and survival; however, structural details of this complex remain elusive. We hypothesize that a conserved, hydrophobic helix (H-anchor) of HKII first inserts into the outer membrane of mitochondria (OMM) and then interacts with VDAC1 on the cytosolic leaflet of OMM to form a binary complex. To systematically investigate this process, we adopted a hybrid approach: 1) the membrane binding of HKII was first described with molecular dynamics (MD) simulations employing a membrane mimetic model with enhanced lipid diffusion, then 2) the resulting membrane-bound HKII was used to form complex with VDAC1 in millisecond-scale Brownian dynamics (BD) simulations. We show that H-anchor inserts its first 10 residues into the membrane, substantiating previous experimental findings. The insertion depth of the H-anchor was used to derive positional restraints in subsequent BD simulations to preserve the membrane-bound pose of HKII during the formation of the HKII/VDAC1 binary complex. Multiple BD-derived structural models were further refined with MD simulations, resulting in one stable complex. A major feature in the complex is the partial (not complete) blockade of VDAC1’s permeation pathway by HKII, a result supported by our comparative electrophysiological measurements of the channel in the presence and absence of HKII. Additionally, we showed how VDAC1 phosphorylation disrupts HKII binding, a feature that is verified by our electrophysiology recordings and have implications in mitochondria-mediated cell death.