DescriptionThe transmembrane pore protein Pannexin-1 (Panx1) forms channels allowing the release of purine nucleotides and participates in processes related to purinergic signaling, including blood pressure regulation, apoptotic cell clearance, and neuropathic pain. Panx1 has also been implicated in several pathological mechanisms, including facilitating HIV entry. While several naturally occurring inhibitors of Panx1 activity have been discovered, elucidating structure-function relationships within the Panx1 oligomer and the ability to design molecules to regulate Panx1 activities has been limited due to the absence of a Panx1 crystal structure. To address this limitation, an in silico model of Panx1 was constructed, based on homologous protein channel structures, established physicochemical characteristics and binding properties of the Panx1 channel.
Homologous template protein searches, transmembrane topologies and sequence alignments were subsequently used by MODELLER software to generate Panx1 A-Chain monomer subunits via a multi-template stitching approach. These A-chain subunit models and a GalaxyWeb Panx1 sequence generated A-chain subunit were subsequently used to construct four model homo-oligomers (pentamers, hexamers, heptamers, and octamers) using GalaxyWEB, all oligomeric structures were compared to established biophysical parameters of Panx1 channels. Docking experiments using Medusa Guide were performed using four small molecules and the 10Panx1 mimetic peptide known to block the Pannexin-1 channel. Docking simulation free energy results were used to assess the regional validity of the designed Panx1 model.
The optimum Pannexin-1 model was a hexameric structure exhibiting compact helices and dynamic loops and tails which promoted both pore and oligomer diameters and an overall structure matching Pannexin-1 experimental data. Docking experiments overall exhibited moderate affinity free energy values for Panx1 inhibitors inferring an overall fair modeling accuracy. A high affinity free energy value of -46.38 kcal mol-1 was discovered for the mimetic 10Panx1 which partially steric blocks the Pannexin-1 pore region.2, 8, 66 EL-1/2 docking results demonstrated high (< -30 to -50 kcal mol-1) to poor (> -20 kcal mol-1) affinities. The model suggested these small molecules bound to the EL1/2 regions primarily through steric/hydrophobic and polar interactions. Overall, this model fits with established experimental results and is a reasonable tool for the initial design of compounds which modulate Pannexin-1 gating.