Analysis of conformational motions and related key residue interactions responsible for a specific function of proteins with elastic network model

Protein collective motions play a critical role in many biochemical processes. How to predict the functional motions and the related key residue interactions in proteins is important for our understanding in the mechanism of the biochemical processes. Normal mode analysis (NMA) of the elastic network model (ENM) is one of the effective approaches to investigate the structure-encoded motions in proteins. However, the motion modes revealed by the conventional NMA approach do not necessarily correspond to a specific function of protein. In the present work, a new analysis method was proposed to identify the motion modes responsible for a specific function of proteins and then predict the key residue interactions involved in the functional motions by using a perturbation approach. In our method, an internal coordinate that accounts for the specific function was introduced, and the Cartesian coordinate space was transformed into the internal/Cartesian space by using linear approximation, where the introduced internal coordinate serves as one of the axes of the coordinate space. NMA of ENM in this internal/Cartesian space was performed and the function-relevant motion modes were identified according to their contributions to the specific function of proteins. Then the key residue interactions important for the functional motions of the protein were predicted as the interactions whose perturbation largely influences the fluctuation along the internal coordinate. Using our proposed methods, the maltose transporter (MalFGK₂) from E. Coli was studied. The functional motions and the key residue interactions that are related to the channel-gating function of this protein were successfully identified.     

Validation of radiocarpal joint contact models based on images from a clinical MRI scanner

This study was undertaken to assess magnetic resonance imaging (MRI)-based radiocarpal surface contact models of functional loading in a clinical MRI scanner for future in vivo studies, by comparison with experimental measures from three cadaver forearm specimens. Experimental data were acquired using a Tekscan sensor during simulated light grasp. Magnetic resonance (MR) images were used to obtain model geometry and kinematics (image registration). Peak contact pressures (PPs) and average contact pressures (APs), contact forces and contact areas were determined in the radiolunate and radioscaphoid joints. Contact area was also measured directly from MR images acquired with load and compared with model data. Based on the validation criteria (within 25% of experimental data), out of the six articulations (three specimens with two articulations each), two met the criterion for AP (0%, 14%); one for peak pressure (20%); one for contact force (5%); four for contact area with respect to experiment (8%, 13%, 19% and 23%), and three contact areas met the criterion with respect to direct measurements (14%, 21% and 21%). Absolute differences between model and experimental PPs were reasonably low (within 2.5 MPa). Overall, the results indicate that MRI-based models generated from 3T clinical MR scanner appear sufficient to obtain clinically relevant data.     

Type-3 functional response in limnetic suspension-feeders,as demonstrated by in situ grazing rates

Field-measured grazing rates (ml/animal/d) of cladocerans (mostly daphniids) and diaptomids were assembled from various published studies and plotted as a function of corresponding phytoplankton concentration (μg l⁻¹ f.w.). Filtering rates of both zooplankton groups initially increased with seston concentration until maximal grazing rates were observed at approximately 4 × 10² and 1 × 10² μg l⁻¹ for cladocerans and copepods, respectively; at higher algal concentrations, filtering rates of both declined as a function of food concentration. The shape of these curves are most consistent with Holling's (1966) Type 3 functional response. We found little support for the Type 3 functional response in published laboratory studies of Daphnia; most investigators report either a Type 1 or Type 2 response. The one study in which the Type 3 response was observed involved experiments where animals were acclimated at low food concentrations for 24 h, whereas those studies associated with response Types 1 or 2 had acclimation periods of only 1 to 3 h. We therefore assembled relevant data from the literature to examine the effect of acclimation period on the feeding rates of Daphnia at low food concentrations. In the absence of any acclimation, animals filtered at extremely low rates. After 2 h of acclimation, however, filtering rates increased 4 to 5-fold but declined again with longer durations; after > 70 h of pre-conditioning, filtering rates were almost as low as they had been with no acclimation. We also found little support for the Type 3 functional response in published studies of copepods. The only study associated with a Type 3 response involved a marine copepod that had been subjected to a starvation period of 48 h; however, an analysis of the effects of acclimation period did not yield conclusive evidence that filtering rates of freshwater copepods (Diaptomus and Eudiaptomus) decrease significantly with acclimation duration. The low filtering rates associated with long acclimation periods in laboratory experiments appears to be a direct result of animals becoming emaciated from prolonged exposure to low food concentrations, a situation which renders them incapable of high filtering rates. This may explain the Type 3 functional response for field cladocerans, since zooplankton in food-limiting situations are constantly exposed to low food concentrations, and would therefore have low body carbon and consequently less energy to filter-feed. We cannot, however, use this to explain the Type 3 response for field diaptomids, since copepods in the laboratory did not appear to lose body carbon even after 72 h of feeding at very low food levels, and there was inconclusive evidence that either Diaptomus or Eudiaptomus decrease their filtering rates with acclimation period. Although Incipient Limiting Concentrations (ILC) for Daphnia ranged from 1 to 8.5 × 10³ μg 1⁻¹, more than half of these fell between 1 and 3 × 10³ μg l⁻¹, bracketing the value of 2.7 × 10² μg l⁻¹ for field cladocerans. There was, however, a great deal of variation in reported maximum ingestion rates (MIR), maximum filtering rates (MFR) and ILC values for Daphnia magna. ILC values from the few laboratory studies of freshwater copepods ranged between 0.5 to 2.8 × 10³ μg 1⁻¹, and was higher than the ILC value of approximately 0.2 × 10³ μg l⁻¹ calculated for field populations of D. minutus. Generally, there was considerable agreement among laboratory studies regarding the shape of grazing-rate and ingestion-rate curves when data were converted to similar units and presented on standardized scales.     

Sensitivity of multifrequency magnetic resonance elastography and diffusion-weighted imaging to cellular and stromal integrity of liver tissue

Microscopic structural alterations of liver tissue induced by freeze-thaw cycles give rise to palpable property changes. However, the underlying damage to tissue architecture is difficult to quantify histologically, and published data on macroscopic changes in biophysical properties are sparse.To better understand the influence of hepatic cells and stroma on global biophysical parameters, we studied rat liver specimens freshly taken (within 30 min after death) and treated by freeze-thaw cycles overnight at either −20 °C or –80 °C using diffusion-weighted imaging (DWI) and multifrequency magnetic resonance elastography (MRE) performed at 0.5 T in a tabletop MRE scanner. Tissue structure was analyzed histologically and rheologic data were analyzed using fractional order derivatives conceptualized by a called spring-pot component that interpolates between pure elastic and viscous responses.Overnight freezing and thawing induced membrane disruptions and cell detachment in the space of Disse, resulting in a markedly lower shear modulus μ and apparent diffusion coefficient (ADC) (μ[−20 °C] = 1.23 ± 0.73 kPa, μ[−80 °C] = 0.66 ± 0.75 kPa; ADC[–20 °C] = 0.649 ± 0.028 μm²/s, ADC[−80 °C] = 0.626 ± 0.025 μm²/s) compared to normal tissue (μ = 9.92 ± 3.30 kPa, ADC = 0.770 ± 0.023 μm²/s, all p < 0.001). Furthermore, we analyzed the springpot-powerlaw coefficient and observed a reduction in −20 °C specimens (0.22 ± 0.14) compared to native tissue (0.40 ± 0.10, p = 0.033) and −80 °C specimens (0.54 ± 0.22, p = 0.002), that correlated with histological observations of sinusoidal dilation and collagen distortion within the space of Disse. Overall, the results suggest that shear modulus and water diffusion in liver tissue markedly decrease due to cell membrane degradation and cell detachment while viscosity-related properties appear to be more sensitive to distorted stromal and microvascular architecture.     

Site-Specific Structural Variations Accompanying Tubular Assembly of the HIV-1 Capsid Protein

The 231-residue capsid (CA) protein of human immunodeficiency virus type 1 (HIV-1) spontaneously self-assembles into tubes with a hexagonal lattice that is believed to mimic the surface lattice of conical capsid cores within intact virions. We report the results of solid-state nuclear magnetic resonance (NMR) measurements on HIV-1 CA tubes that provide new information regarding changes in molecular structure that accompany CA self-assembly, local dynamics within CA tubes, and possible mechanisms for the generation of lattice curvature. This information is contained in site-specific assignments of signals in two- and three-dimensional solid-state NMR spectra, conformation-dependent ¹⁵N and ¹³C NMR chemical shifts, detection of highly dynamic residues under solution NMR conditions, measurements of local variations in transverse spin relaxation rates of amide ¹H nuclei, and quantitative measurements of site-specific ¹⁵N–¹⁵N dipole–dipole couplings. Our data show that most of the CA sequence is conformationally ordered and relatively rigid in tubular assemblies and that structures of the N-terminal domain (NTD) and the C-terminal domain (CTD) observed in solution are largely retained. However, specific segments, including the N-terminal β-hairpin, the cyclophilin A binding loop, the inter-domain linker, segments involved in intermolecular NTD–CTD interactions, and the C-terminal tail, have substantial static or dynamical disorder in tubular assemblies. Other segments, including the 3₁₀-helical segment in CTD, undergo clear conformational changes. Structural variations associated with curvature of the CA lattice appear to be localized in the inter-domain linker and intermolecular NTD–CTD interface, while structural variations within NTD hexamers, around local 3-fold symmetry axes, and in CTD–CTD dimerization interfaces are less significant.