No abstract

Cell membranes contain a large variety of lipid types and are crowded with proteins, endowing them with the plasticity needed to fulfill their key roles in cell functioning. The compositional complexity of cellular membranes gives rise to a heterogeneous lateral organization, which is still poorly understood. Computational models, in particular molecular dynamics simulations and related techniques, have provided important insight into the organizational principles of cell membranes over the past decades. Now, we are witnessing a transition from simulations of simpler membrane models to multicomponent systems, culminating in realistic models of an increasing variety of cell types and organelles. Here, we review the state of the art in the field of realistic membrane simulations and discuss the current limitations and challenges ahead.

Developmental signals of the Hedgehog (Hh) and Wnt families are transduced across the membrane by Frizzled-class G-protein coupled receptors (GPCRs) composed of both a heptahelical transmembrane domain (TMD) and an extracellular cysteine-rich domain (CRD). How such large extracellular domains of GPCRs regulate signalling by the TMD is unknown. We present crystal structures of the Hh signal transducer and oncoprotein Smoothened (SMO), which contains two distinct ligand-binding sites in its TMD and CRD. The CRD is stacked atop the TMD, separated by an intervening wedge-like linker domain (LD). Structure-guided mutations show that the interface between the CRD, LD and TMD stabilises the inactive state of SMO. Unexpectedly, we find a cholesterol molecule bound to SMO in the CRD-binding site. Mutations predicted to prevent cholesterol binding impair the ability of SMO to transmit native Hh signals. Binding of a clinically used antagonist, vismodegib, to the TMD induces a conformational change that is propagated to the CRD, resulting in loss of cholesterol from the CRD-LD-TMD interface. Our work elucidates the structural mechanism by which the activity of a GPCR is controlled by ligand-regulated interactions between its extracellular and transmembrane domains.

Water plays a key role in biological membrane transport. In ion channels and water-conducting pores (aquaporins), one-dimensional confinement in conjunction with strong surface effects changes the physical behavior of water. In molecular dynamics simulations of water in short (0.8 nm) hydrophobic pores the water density in the pore fluctuates on a nanosecond time scale. In long simulations (460 ns in total) at pore radii ranging from 0.35 to 1.0 nm we quantify the kinetics of oscillations between a liquid-filled and a vapor-filled pore. This behavior can be explained as capillary evaporation alternating with capillary condensation, driven by pressure fluctuations in the water outside the pore. The freeenergy difference between the two states depends linearly on the radius. The free-energy landscape shows how a metastable liquid state gradually develops with increasing radius. For radii > Ϸ0.55 nm it becomes the globally stable state and the vapor state vanishes. One-dimensional confinement affects the dynamic behavior of the water molecules and increases the self diffusion by a factor of 2-3 compared with bulk water. Permeabilities for the narrow pores are of the same order of magnitude as for biological water pores. Water flow is not continuous but occurs in bursts. Our results suggest that simulations aimed at collective phenomena such as hydrophobic effects may require simulation times >50 ns. For water in confined geometries, it is not possible to extrapolate from bulk or short time behavior to longer time scales. C hannel and transporter proteins control flow of water, ions, and other solutes across cell membranes. In recent years several channel and pore structures have been solved at near atomic resolution (1-6), which together with three decades of physiological data (7) and theoretical and simulation approaches (8) allow us to describe transport of ions, water, or other small molecules at a molecular level. Water plays a special role here: it either solvates the inner surfaces of the pore and the permeators (for example, ions and small molecules like glycerol), or it is the permeant species itself as in the aquaporin family of water pores (9-11) or in the bacterial peptide channel gramicidin A, whose water transport properties are well studied (12-14). Thus, a better characterization of the behavior of water would improve our understanding of the biological function of a wide range of transporters. The remarkable water transport properties of aquaporins [water is conducted through a long (Ϸ2 nm) and narrow (Ϸ0.3 nm diameter) pore at bulk diffusion rates while at the same time protons are strongly selected against] are the topic of recent simulation studies (15, 16).The shape and dimensions of biological pores and the nature of the pore-lining atoms are recognized as major determinants of function. How the behavior of water depends on these factors is far from understood (17). Water is not a simple liquid because of its strong hydrogen bond network. When confined to narrow geometries like slits or pores ...