Structures of human pannexin-1 in nanodiscs reveal gating mediated by dynamic movement of the N terminus and phospholipids.

Pannexin (PANX) family proteins form large-pore channels that mediate purinergic signaling. We analyzed the cryo-EM structures of human PANX1 in lipid nanodiscs to elucidate the gating mechanism and its regulation by the amino terminus in phospholipids. The wild-type channel has an amino-terminal funnel in the pore, but in the presence of the inhibitor probenecid, a cytoplasmically oriented amino terminus and phospholipids obstruct the pore. Functional analysis using whole-cell patch-clamp and oocyte voltage clamp showed that PANX1 lacking the amino terminus did not open and had a dominant negative effect on channel activity, thus confirming that the amino-terminal domain played an essential role in channel opening. These observations suggest that dynamic conformational changes in the amino terminus of human PANX1 are associated with lipid movement in and out of the pore. Moreover, the data provide insight into the gating mechanism of PANX1 and, more broadly, other large-pore channels.

Optimizing Go-MARTINI Coarse-Grained Model for F-BAR Protein on Lipid Membrane.

Coarse-grained (CG) molecular dynamics (MD) simulations allow us to access much larger length and time scales than atomistic MD simulations, providing an attractive alternative to the conventional simulations. Based on the well-known MARTINI CG force field, the recently developed Go-MARTINI model for proteins describes large-amplitude structural dynamics, which has not been possible with the commonly used elastic network model. Using the Go-MARTINI model, we conduct MD simulations of the F-BAR Pacsin1 protein on lipid membrane. We observe that structural changes of the non-globular protein are largely dependent on the definition of the native contacts in the Go model. To address this issue, we introduced a simple cutoff scheme and tuned the cutoff distance of the native contacts and the interaction strength of the Lennard-Jones potentials in the Go-MARTINI model. With the optimized Go-MARTINI model, we show that it reproduces structural fluctuations of the Pacsin1 dimer from atomistic simulations. We also show that two Pacsin1 dimers properly assemble through lateral interaction on the lipid membrane. Our work presents a first step towards describing membrane remodeling processes in the Go-MARTINI CG framework by simulating a crucial step of protein assembly on the membrane.

Curvature induction and sensing of the F-BAR protein Pacsin1 on lipid membranes via molecular dynamics simulations.

F-Bin/Amphiphysin/Rvs (F-BAR) domain proteins play essential roles in biological processes that involve membrane remodelling, such as endocytosis and exocytosis. It has been shown that such proteins transform the lipid membrane into tubes. Notably, Pacsin1 from the Pacsin/Syndapin subfamily has the ability to transform the membrane into various morphologies: striated tubes, featureless wide and thin tubes, and pearling vesicles. The molecular mechanism of this interesting ability remains elusive. In this study, we performed all-atom (AA) and coarse-grained (CG) molecular dynamics simulations to investigate the curvature induction and sensing mechanisms of Pacsin1 on a membrane. From AA simulations, we show that Pacsin1 has internal structural flexibility. In CG simulations with parameters tuned from the AA simulations, spontaneous assembly of two Pacsin1 dimers through lateral interaction is observed. Based on the complex structure, we show that the regularly assembled Pacsin1 dimers bend a tensionless membrane. We also show that a single Pacsin1 dimer senses the membrane curvature, binding to a buckled membrane with a preferred curvature. These results provide molecular insights into polymorphic membrane remodelling.

Binding and aggregation mechanism of amyloid ß-peptides onto the GM1 ganglioside-containing lipid membrane.

Accumulation and fibril formation of amyloid ß (Aß) peptides onto a ganglioside-rich lipid membrane is a cause of neuro-disturbance diseases. To find out a measure for suppressing the nucleation of a seed for amyloid fibrils, the mechanism of the initial binding of Aß to the membrane should be clarified. Molecular dynamics simulations were carried out to investigate the adhesion process of Aß peptides onto a GM1-ganglioside-containing membrane. Multiple computational trials were executed to analyze the probability of occurrence of Aß binding by using calculation models containing a mixed lipid membrane, water layer, and one, two, or three Aßs. The simulations demonstrated that Aß peptides approached the membrane after fluctuation in the water layer and occasionally made steady contact with the membrane. Once the steady contact had been established, Aß was unlikely to be detached from the membrane and developed into a more stably bound form. In the stably bound form, neuraminic acids on the GM1 cluster strongly held the side chain of Lys28 of Aß, which caused deformation of the C-terminal region of the Aß. Since the C-terminal region of the Aß peptide contains many hydrophobic residues, its deformation on the membrane enhances the hydrophobic interaction with other Aß peptides. The contact region of two Aßs evolved into a parallel ß-sheet form, and the third Aß was observed to be bound to the complex of two Aßs to make a bundle of Aß peptides. Some key structures involved in the Aß aggregation on the GM1-containing membrane were deduced from the multiple simulations.

Formation of GM1 ganglioside clusters on the lipid membrane containing sphingomyeline and cholesterol.

GM1 gangliosides form a microdomain with sphingomyeline (SM) and cholesterol (Chol) and are deeply involved in the aggregation of amyloid beta (Aß) peptides on neural membranes. We performed molecular dynamics simulations on two kinds of lipid bilayers containing GM1 ganglioside: GM1/SM/Chol and GM1/POPC. Both 10 and 100 ns simulations and another set of 10 ns simulations with different initial lipid arrangement essentially showed the same computational results. GM1 molecules in the GM1/SM/Chol membrane were condensed, whereas those in GM1/POPC membrane scattered. That is, the formation of GM1 cluster was observed only on the GM1/SM/Chol mixed membrane. There appeared numerous hydrogen bonds among glycan portions of the GM1 clusters due to the condensation. A comparison in distribution of lipid molecules between the two kinds of membranes suggested that cholesterol had important roles to prevent the membrane from interdigitation and to stabilize other lipids for interacting with each other. This property of cholesterol promotes the formation of GM1 clusters.