Unlike dilute experimental conditions under which biological molecules are typically characterized, the cell interior is crowded by macromolecules, which affects both the thermodynamics and kinetics ...of in vivo processes. Although the excluded-volume effects of macromolecular crowding are expected to cause compaction of unfolded and disordered proteins, the extent of this effect is uncertain. We use a coarse-grained model to represent proteins with varying sequence content and directly observe changes in chain dimensions in the presence of purely repulsive spherical crowders. We find that the extent of crowding-induced compaction is dependent not only on crowder size and concentration, but also on the properties of the protein itself. In fact, we observe a nonmonotonic trend between the dimensions of the polypeptide chain in bulk and the degree of compaction: the most extended chains experience up to 24% compaction, the most compact chains show virtually no change, and intermediate chains compress by up to 40% in size at a 40% crowder volume fraction. Free-volume theory combined with an impenetrable ellipsoidal representation of the chains predicts the crowding effects only for collapsed protein chains. An additional scaling factor, which can be easily computed from protein-crowder potential of mean force, corrects for the penetrability of extended chains and is sufficient to capture the observed nonmonotonic trend in compaction.
Abstract
Dynamic remodeling of the actin cytoskeleton is essential for many cellular processes. Tracking the movement of individual actin filaments can in principle shed light on how this complex ...behavior arises at the molecular level. However, the information that can be extracted from these measurements is often limited by low signal-to-noise ratios. We developed a Bayesian statistical approach to estimate true, underlying velocity distributions from the tracks of individual actin-associated fluorophores with quantified localization uncertainties. We found that the motion of filamentous (F)-actin in fibroblasts and endothelial cells was better described by a statistical jump process than by models in which filaments undergo continuous, diffusive movement. In particular, a model with exponentially distributed jump length- and time-scales recapitulated actin filament velocity distributions measured for the cell cortex, integrin-based adhesions, and stress fibers, suggesting that a common physical model can potentially describe actin filament dynamics in a variety of cellular contexts.
Intrinsically disordered proteins (IDPs), which are expected to be largely unstructured under physiological conditions, make up a large fraction of eukaryotic proteins. Molecular dynamics simulations ...have been utilized to probe structural characteristics of these proteins, which are not always easily accessible to experiments. However, exploration of the conformational space by brute force molecular dynamics simulations is often limited by short time scales. Present literature provides a number of enhanced sampling methods to explore protein conformational space in molecular simulations more efficiently. In this work, we present a comparison of two enhanced sampling methods: temperature replica exchange molecular dynamics and bias exchange metadynamics. By investigating both the free energy landscape as a function of pertinent order parameters and the per-residue secondary structures of an IDP, namely, human islet amyloid polypeptide, we found that the two methods yield similar results as expected. We also highlight the practical difference between the two methods by describing the path that we followed to obtain both sets of data.
The cholesterol recognition/interaction amino acid consensus (CRAC) motif is a primary structure pattern used to identify regions that may be responsible for preferential cholesterol binding in many ...proteins. The leukotoxin LtxA, which is produced by a pathogenic bacterium, contains two CRAC seqences, only one of which is responsible for cholesterol binding, and the binding is required for cytotoxicity. The factors, in addition to the CRAC definition, that may be responsible for cholesterol-binding functionality and atomistic interactions between the CRAC region and cholesterol are as yet unknown. This study uses molecular dynamics simulations to identify structural characteristics and specific interactions of the two LtxA CRAC peptides with both pure phospholipid and binary cholesterol–phospholipid bilayers. We have identified changes in the secondary structure of these peptides that occur upon cholesterol binding, which are not seen when it is associated with a cholesterol-devoid membrane, and which show salient coupling of structural disorder and function. Additionally, the central tyrosine residue of the CRAC motif was found to play a significant role in cholesterol binding, though residues outside of the CRAC motif also influence membrane interactions and functionality of the CRAC region.
Role of solvation in pressure-induced helix stabilization Best, Robert B; Miller, Cayla; Mittal, Jeetain
Journal of chemical physics online/The Journal of chemical physics/Journal of chemical physics,
12/2014, Letnik:
141, Številka:
22
Journal Article
Recenzirano
Odprti dostop
In contrast to the well-known destabilization of globular proteins by high pressure, recent work has shown that pressure stabilizes the formation of isolated α-helices. However, all simulations to ...date have obtained a qualitatively opposite result within the experimental pressure range. We show that using a protein force field (Amber03w) parametrized in conjunction with an accurate water model (TIP4P/2005) recovers the correct pressure-dependence and an overall stability diagram for helix formation similar to that from experiment; on the other hand, we confirm that using TIP3P water results in a very weak pressure destabilization of helices. By carefully analyzing the contributing factors, we show that this is not merely a consequence of different peptide conformations sampled using TIP3P. Rather, there is a critical role for the solvent itself in determining the dependence of total system volume (peptide and solvent) on helix content. Helical peptide structures exclude a smaller volume to water, relative to non-helical structures with both the water models, but the total system volume for helical conformations is higher than non-helical conformations with TIP3P water at low to intermediate pressures, in contrast to TIP4P/2005 water. Our results further emphasize the importance of using an accurate water model to study protein folding under conditions away from standard temperature and pressure.
Integrin-based adhesion complexes link the cytoskeleton to the extracellular matrix (ECM) and are central to the construction of multicellular animal tissues. How biological function emerges from the ...tens to thousands of proteins present within a single adhesion complex remains unclear. We used fluorescent molecular tension sensors to visualize force transmission by individual integrins in living cells. These measurements revealed an underlying functional modularity in which integrin class controlled adhesion size and ECM ligand specificity, while the number and type of connections between integrins and F-actin determined the force per individual integrin. In addition, we found that most integrins existed in a state of near-mechanical equilibrium, a result not predicted by existing models of cytoskeletal force transduction. A revised model that includes reversible cross-links within the F-actin network can account for this result and suggests one means by which cellular mechanical homeostasis can arise at the molecular level.
Hearing depends on precise synaptic transmission between cochlear inner hair cells and spiral ganglion neurons through afferent ribbon synapses.
Neuroligins (Nlgns) facilitate synapse maturation in ...the brain, but they have gone unstudied in the cochlea. We report Nlgn3 and Nlgn1 knockout (KO) cochleae have fewer ribbon synapses and have impaired hearing. Nlgn3 KO is more vulnerable to noise trauma with limited activity at high frequencies one day after noise. Furthermore, Nlgn3 KO cochleae have a 5-fold reduction in synapse number compared to wild type after two weeks of recovery. Double KO cochlear phenotypes are more prominent than the KOs, for example, 5-fold smaller synapses, 25% reduction in synapse density, and 30% less synaptic output. These observations indicate Nlgn3 and Nlgn1 are essential to cochlear ribbon synapse maturation and function.
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•Nlgn3 is a major synapse organizing protein in spiral ganglion neurons•Nlgn1 and three KO cochlea have reduced ribbon synapse density and hampered function•Nlgn3 KO mice are highly sensitive to noise exposures•dKO cochlear phenotypes are generally synergistic and severe
Genomics; Neuroscience; Cellular neuroscience; Sensory neuroscience
Human islet amyloid polypeptide (hIAPP), a 37-residue protein cosecreted with insulin by β-cells in the pancreas, is known to form amyloid fibrils in type II diabetes patients. This fibril formation ...is also associated with β-cell death. However, rat IAPP (rIAPP) does not aggregate into fibrils, nor is it associated with β-cell toxicity. Determining solution properties of hIAPP experimentally is difficult because it aggregates quickly. Our study uses molecular dynamics simulation to explore and compare in-solution characteristics of hIAPP and rIAPP, as well as two single-point hIAPP mutants, hIAPP I26P and hIAPP S20G, which exhibit observed differences from hIAPP in aggregation propensities. We find that all four polypeptide monomers sample structured states in solution. More importantly, differences in the helicity over residues 7–16 may play an important role in early aggregation, although this region is outside of commonly assumed amyloidogenic region 20–29. The long-range contacts, though unexpected of IDPs, cause variation in sampled conformations among four polypeptides within same amino acid sequence. Our results also yield evidence that previously determined structures bound to micelles are also transiently sampled in the solution state. In particular, similarities found in region 8–17 together with the helical differences that we observe in region 7–16 lead us to suggest that the region 7–16 is potentially responsible for amyloidogenic behavior of amylin peptides. Our results also provide support for the proposed mechanism of fibril formation based on experimentally observed transient helices in amyloidogenic peptides.
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