Phosphatidylserine (PS) is a negatively charged lipid type commonly found in eukaryotic membranes, where it interacts with proteins via nonspecific electrostatic interactions as well as via specific binding. Moreover, in the presence of calcium ions, PS lipids can induce membrane fusion and phase separation. Molecular details of these phenomena remain poorly understood, partly because accurate models to interpret the experimental data have not been available. Here we gather a set of previously published experimental NMR data of C-H bond order parameter magnitudes, |S CH |, for pure PS and mixed PS:PC (phosphatidylcholine) lipid bilayers, and augment this data set by measuring the signs of S CH in the PS headgroup using S-DROSS solid-state NMR spectroscopy. The augmented data set is then used to assess the accuracy of the PS headgroup structures in, and the cation binding to, PS-containing membranes in the most commonly used classical molecular dynamics (MD) force fields including CHARMM36, Lipid17, MacRog, Slipids, GROMOS-CKP, Berger, and variants. We show large discrepancies between different force fields, and that none of them reproduces the NMR data within experimental accuracy. However, the best MD models can detect the most essential differences between PC and PS headgroup structures. The cation binding affinity is, in line with our previous results for PC lipids, not captured correctly by any of the PS force fields. Moreover, the simulated response of PS headgroup to bound ions can differ from experiments even qualitatively. The collected experimental dataset and simulation results will pave the way for development of lipid force fields that correctly describe the biologically relevant negatively charged membranes and their interactions with ions. This work is part of the NMRlipids open collaboration project (nmrlipids.blogspot.fi).

The progress towards understanding the molecular basis of Alzheimers's disease is strongly connected to elucidating the early aggregation events of the amyloid-β (Aβ) peptide. Molecular dynamics (MD) simulations provide a viable technique to study the aggregation of Aβ into oligomers with high spatial and temporal resolution. However, the results of an MD simulation can only be as good as the underlying force field. A recent study by our group showed that none of the force fields tested can distinguish between aggregation-prone and non-aggregating peptide sequences, producing the same and in most cases too fast aggregation kinetics for all peptides. Since then, new force fields specially designed for intrinsically disordered proteins such as Aβ were developed. Here, we assess the applicability of these new force fields to studying peptide aggregation using the Aβ 16−22 peptide and mutations of it as test case. We investigate their performance in modeling the monomeric state, the aggregation into oligomers, and the stability of the aggregation end product, i.e., the fibrillar state. A main finding is that changing the force field has a stronger effect on the simulated aggregation pathway than changing the peptide sequence. Also the new force fields are not able to reproduce the experimental aggregation propensity order of the peptides. Dissecting the various energy contributions shows that AMBER99SB-disp overestimates the interactions between the peptides and water, thereby inhibiting peptide aggregation.More promising results are obtained with CHARMM36m and especially its version with increased protein-water interactions. It is thus recommended to use this force field for peptide aggregation simulations and base future reparameterizations on it. IntroductionIntrinsically disordered proteins (IDPs) are a class of proteins which are either completely unstructured or contain large disordered regions in their native state, which comes with a high tendency towards self-assembly leading to non-toxic or toxic aggregates and fibrils that may be related to diseases. 1 One of the IDPs is the amyloid-beta peptide (Aβ), forming 2

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The generation of mechanical forces are central to a wide range of vital biological processes, including the function of the cytoskeleton. Although the forces emerging from the polymerization of native proteins have been studied in detail, the potential for force generation by aberrant protein polymerization has not yet been explored. Here, we show that the growth of amyloid fibrils, archetypical aberrant protein polymers, is capable of unleashing mechanical forces on the piconewton scale for individual filaments. We apply microfluidic techniques to measure the forces released by amyloid growth for two systems: insulin and lysozyme. The level of force measured for amyloid growth in both systems is comparable to that observed for actin and tubulin, systems that have evolved to generate force during their native functions and, unlike amyloid growth, rely on the input of external energy in the form of nucleotide hydrolysis for maximum force generation. Furthermore, we find that the power density released from growing amyloid fibrils is comparable to that of high-performance synthetic polymer actuators. These findings highlight the potential of amyloid structures as active materials and shed light on the criteria for regulation and reversibility that guide molecular evolution of functional polymers.microfluidics | amyloidosis | active materials | biological force generation | protein misfolding

Carbohydrates such as the trisaccharide motif LeX are key constituents of cell surfaces. Despite intense research, the interactions between carbohydrates of apposing cells or membranes are not well understood. In...