The influence of parameters of consecutive speed control humps ...

This paper is aimed at analyzing the chaotic vibration of a vehicle passing the consecutive speed control humps (SCHs) on a highway. A consecutive SCHs-speed coupling excitation function is presented. The chaotic vibration of nonlinear vehicle is studied by numerical simulation under a 2-DOF nonlinear vehicle suspension model. The chaotic vibration excited by the consecutive SCHs with different parameters is analyzed. Simulation results demonstrate that the chaotic motion may occur as the vehicle moves over a series of the consecutive SCHs. Furthermore, chaotic motion can be inhibited reasonably and effectively by proper adjustment of parameters of the consecutive SCHs

Using NMR to Distinguish Viscosity Effects from Nonspecific Protein Binding under Crowded Conditions

Conventional NMR approaches to detect weak protein binding and aggregation are hindered by the increased viscosity brought about by crowding. We describe a simple and reliable NMR method to distinguish viscosity effects from binding and aggregation under crowded conditions

Effects of Proteins on Protein Diffusion

Despite increased attention, little is known about how the crowded intracellular environment affects basic phenomena like protein diffusion. Here, we use NMR to quantify the rotational and translational diffusion of a 7.4-kDa test protein, chymotrypsin inhibitor 2 (CI2), in solutions of glycerol, synthetic polymers, proteins, and cell lysates. As expected, translational diffusion and rotational diffusion decrease with increasing viscosity. In glycerol, for example, the decrease follows the Stokes-Einstein and Stokes-Einstein-Debye laws. Synthetic polymers cause negative deviation from the Stokes Laws and affect translation more than rotation. Surprisingly, however, protein crowders have the opposite effect, causing positive deviation and reducing rotational diffusion more than translational diffusion. Indeed, bulk proteins severely attenuate the rotational diffusion of CI2 in crowded protein solutions. Similarly, CI2 diffusion in cell lysates is comparable to its diffusion in crowded protein solutions, supporting the biological relevance of the results. The rotational attenuation is independent of the size and total charge of the crowding protein, suggesting that the effect is general. The difference between the behavior of synthetic polymers and protein crowders suggests that synthetic polymers may not be suitable mimics of the intracellular environment. NMR relaxation data reveal that the source of the difference between synthetic polymers and proteins is the presence of weak interactions between the proteins and CI2. In summary, weak but non-specific, non-covalent chemical interactions between proteins appear to fundamentally impact protein diffusion in cells

19 F NMR studies of α-synuclein-membrane interactions: α-Synuclein-Membrane Interactions

α-Synuclein function is thought to be related to its membrane binding ability. Solution NMR studies have identified several α-synuclein-membrane interaction modes in small unilamellar vesicles (SUVs), but how membrane properties affect binding remains unclear. Here, we use 19F NMR to study α-synuclein-membrane interactions by using 3-fluoro-L-tyrosine (3FY) and trifluoromethyl-L-phenylalanine (tfmF) labeled proteins. Our results indicate that the affinity is affected by both the head group and the acyl chain of the SUV. Negatively charged head groups have higher affinity, but different head groups with the same charge also affect binding. We show that the saturation of the acyl chain has a dramatic effect on the α-synuclein-membrane interactions by studying lipids with the same head group but different chains. Taken together, the data show that α-synuclein's N-terminal region is the most important determinate of SUV binding, but its C-terminal region also modulates the interactions. Our data support the existence of multiple tight phospholipid-binding modes, a result incompatible with the model that α-synuclein lies solely on the membrane surface

Translational and Rotational Diffusion of a Small Globular Protein under Crowded Conditions

Protein protein interaction is the fundamental step of biological signal transduction. Interacting proteins find each other by diffusion. To gain insight into diffusion under the crowded conditions found in cells, we used nuclear magnetic resonance spectroscopy (NMR) to measure the effects of solvent additives on the translational and rotational diffusion of the 7.4 kDa globular protein, chymotrypsin inhibitor 2. The additives were glycerol and the macromolecular crowding agent, polyvinylpyrrolidone (PVP). Both translational diffusion and rotational diffusion decrease with increasing solution viscosity. For glycerol, the decrease obeys the Stokes Einstein and Stokes Einstein Debye laws. Three types of deviation are observed for PVP: the decrease in diffusion with increased viscosity is less than predicted, this negative deviation is greater for rotational diffusion, and the negative deviation increases with increasing PVP molecular weight. We discuss our results in terms of other studies on the effects of macromolecules on globular protein diffusion

An upper limit for macromolecular crowding effects

<p>Abstract</p> <p>Background</p> <p>Solutions containing high macromolecule concentrations are predicted to affect a number of protein properties compared to those properties in dilute solution. In cells, these macromolecular crowders have a large range of sizes and can occupy 30% or more of the available volume. We chose to study the stability and ps-ns internal dynamics of a globular protein whose radius is ~2 nm when crowded by a synthetic microgel composed of poly(<it>N</it>-isopropylacrylamide-<it>co</it>-acrylic acid) with particle radii of ~300 nm.</p> <p>Results</p> <p>Our studies revealed no change in protein rotational or ps-ns backbone dynamics and only mild (~0.5 kcal/mol at 37°C, pH 5.4) stabilization at a volume occupancy of 70%, which approaches the occupancy of closely packing spheres. The lack of change in rotational dynamics indicates the absence of strong crowder-protein interactions.</p> <p>Conclusions</p> <p>Our observations are explained by the large size discrepancy between the protein and crowders and by the internal structure of the microgels, which provide interstitial spaces and internal pores where the protein can exist in a dilute solution-like environment. In summary, microgels that interact weakly with proteins do not strongly influence protein dynamics or stability because these large microgels constitute an upper size limit on crowding effects.</p