Styrene-Maleic Acid Copolymer Nanodiscs to Determine the Shape of Membrane Proteins.

Lipid nanodiscs can be used to solubilize functional membrane proteins (MPs) in nativelike environments. Thus, they are promising reagents that have been proven useful to characterize MPs. Both protein and non-protein molecular belts have shown promise to maintain the structural integrity of MPs in lipid nanodiscs. Small-angle neutron scattering (SANS) can be used to determine low-resolution structures of proteins in solution, which can be enhanced through the use of contrast variation methods. We present theoretical contrast variation SANS results for protein and styrene-maleic acid copolymer (SMA) belt 1,2-dimyristoyl-sn-glycero-3-phosphorylcholine (DMPC) nanodiscs with and without additional bound or transmembrane proteins. The predicted scattering properties are derived from atomistic molecular dynamics simulations to account for conformational fluctuations, and we determine deuterium-labeling conditions such that SANS intensity profiles only include contributions from the scattering of the MP of interest. We propose strategies to tune the neutron scattering length densities (SLDs) of the SMA and DMPC using selective deuterium labeling such that the SLD of the nanodisc becomes homogeneous and its scattering can essentially be eliminated in solvents containing an appropriate amount of D2O. These finely tuned labeled polymer-based nanodiscs are expected to be useful to extract the size and molecular shape information of MPs using SANS-based contrast variation experiments, and they can be used with MPs of any molecular weight.

Kinetic CRAC uncovers a role for Nab3 in determining gene expression profiles during stress.

RNA-binding proteins play a key role in shaping gene expression profiles during stress, however, little is known about the dynamic nature of these interactions and how this influences the kinetics of gene expression. To address this, we developed kinetic cross-linking and analysis of cDNAs (χCRAC), an ultraviolet cross-linking method that enabled us to quantitatively measure the dynamics of protein-RNA interactions in vivo on a minute time-scale. Here, using χCRAC we measure the global RNA-binding dynamics of the yeast transcription termination factor Nab3 in response to glucose starvation. These measurements reveal rapid changes in protein-RNA interactions within 1 min following stress imposition. Changes in Nab3 binding are largely independent of alterations in transcription rate during the early stages of stress response, indicating orthogonal transcriptional control mechanisms. We also uncover a function for Nab3 in dampening expression of stress-responsive genes. χCRAC has the potential to greatly enhance our understanding of in vivo dynamics of protein-RNA interactions.Protein RNA interactions are dynamic and regulated in response to environmental changes. Here the authors describe 'kinetic CRAC', an approach that allows time resolved analyses of protein RNA interactions with minute time point resolution and apply it to gain insight into the function of the RNA-binding protein Nab3.

Load-induced stresses in photoelastic primary canines with facial restorations.

PURPOSE: The purpose of this study was to photoelastically compare the stresses generated by loads on primary canines with facial restorations of different stiffness. METHODS: Composite photoelastic models of a typical maxillary primary canine were fabricated using individual simulant materials for enamel, dentin, periodontal ligament, and alveolar bone. Models were made with identical facial preparations included either near the cementoenamel junction or at 2 mm incisally. The model teeth were restored using the following materials of disparate elastic moduli: (1) high modulus, hybrid composite (Herculite XRV); and (2) lower modulus compomer (Dyract). Three replications of each type of restored tooth were fabricated. Simulated masticatory forces were applied on the cusp tip and the cingulum of each tooth model. The resulting stress patterns were observed and recorded photographically in the field of a circular polariscope. RESULTS: Prior to load application, similar low-level, polymerization-induced shrinkage stresses were localized at the preparation margins of all models. Under both incisal and cingular loading, the higher modulus hybrid composite tended to concentrate stress along the gingival and proximal margins more than did the lower modulus compomer. This effect was more pronounced with cingular than with incisal loading. For both preparations and restoratives, higher stresses were produced by the cingulum loading. No significant differences were observed within each group of replicated restored models. CONCLUSION: These results suggest that, regardless of preparation height, lower modulus compomer restoration of facial lesions in primary maxillary canines may reduce stress production by occlusal forces.