Concentrating membrane proteins using asymmetric traps and AC electric fields

Membrane proteins are key components of the plasma membrane and are responsible for control of chemical ionic gradients, metabolite and nutrient transfer, and signal transduction between the interior of cells and the external environment. Of the genes in the human genome, 30% code for membrane proteins (Krogh et al. J. Mol. Biol.2001, 305, 567). Furthermore, many FDA-approved drugs target such proteins (Overington et al. Nat. Rev. Drug Discovery 2006, 5, 993). However, the structure-function relationships of these are notably sparse because of difficulties in their purification and handling outside of their membranous environment. Methods that permit the manipulation of membrane components while they are still in the membrane would find widespread application in separation, purification, and eventual structure-function determination of these species (Poo et al. Nature 1977, 265, 602). Here we show that asymmetrically patterned supported lipid bilayers in combination with AC electric fields can lead to efficient manipulation of charged components. We demonstrate the concentration and trapping of such components through the use of a "nested trap" and show that this method is capable of yielding an approximately 30-fold increase in the average protein concentration. Upon removal of the field, the material remains trapped for several hours as a result of topographically restricted diffusion. Our results indicate that this method can be used for concentrating and trapping charged membrane components while they are still within their membranous environment. We anticipate that our approach could find widespread application in the manipulation and study of membrane proteins.     

Observation of the charge symmetry breaking d+d-->4He+pi(0) reaction near threshold

We report the first observation of the charge symmetry breaking d+d-->4He+pi(0) reaction near threshold. Measurements using a magnetic channel (gated by two photons) of the 4He scattering angle and momentum (from time of flight) permitted reconstruction of the pi(0) "missing mass," the quantity used to separate 4He+pi(0) events from the continuum of double radiative capture 4He+gamma+gamma events. We measured total cross sections for neutral pion production of 12.7+/-2.2 pb at 228.5 MeV and 15.1+/-3.1 pb at 231.8 MeV. The uncertainty is dominated by statistical errors. These cross sections arise fundamentally from the down-up quark mass difference and quark electromagnetic effects that contribute in part through meson mixing (e.g., pi(0)-eta) mechanisms.     

Absolute stereochemical determination of chiral carboxylic acids

Monoamidation of 1,4-diaminobenzene with alpha-chiral carboxylic acids leads to a carrier strategy for absolute stereochemical determination with bis-zinc porphyrin tweezers by exciton-coupled circular dichroism (ECCD). The helicity induced in the porphyrin tweezers upon complexation with the derivatized carrier originates from the preferred conformation of the C(carbonyl)-C(chiral) bond. Correct ECCD signs can be predicted by the rotamer that places the large group perpendicular to the carbonyl group with the small group facing the porphyrin. [reaction: see text]     

NMR Crystallography: Evaluation of Hydrogen Positions in Hydromagnesite by 13C{1H} REDOR Solid‐State NMR and Density Functional Theory Calculation of Chemical Shielding Tensors

Solid‐state NMR measurements coupled with density functional theory (DFT) calculations demonstrate how hydrogen positions can be refined in a crystalline system. The precision afforded by rotational‐echo double‐resonance (REDOR) NMR to interrogate ¹³C–¹H distances is exploited along with DFT determinations of the ¹³C tensor of carbonates (CO₃^2?). Nearby ¹H nuclei perturb the axial symmetry of the carbonate sites in the hydrated carbonate mineral, hydromagnesite [4 MgCO₃?Mg(OH)₂?4 H₂O]. A match between the calculated structure and solid‐state NMR was found by testing multiple semi‐local and dispersion‐corrected DFT functionals and applying them to optimize atom positions, starting from X‐ray diffraction (XRD)‐determined atomic coordinates. This was validated by comparing calculated to experimental ¹³C{¹H} REDOR and ¹³C chemical shift anisotropy (CSA) tensor values. The results show that the combination of solid‐state NMR, XRD, and DFT can improve structure refinement for hydrated materials.     

Nonlinear dynamics of an interface between shear bands

We study numerically the nonlinear dynamics of a shear banding interface in two-dimensional planar shear flow, within the nonlocal Johnson-Segalman model. Consistent with a recent linear stability analysis, we find that an initially flat interface is unstable with respect to small undulations for a sufficiently small ratio of the interfacial width l to cell length L(x). The instability saturates in finite amplitude interfacial fluctuations. For decreasing l/L(x) these undergo a nonequilibrium transition from simple traveling interfacial waves with constant average wall stress, to periodically rippling waves with a periodic stress response. When multiple shear bands are present we find erratic interfacial dynamics and a stress response suggesting low dimensional chaos.     

A minimal model for vorticity and gradient banding in complex fluids

A general phenomenological reaction-diffusion model for flow-induced phase transitions in complex fluids is presented. The model consists of an equation of motion for a nonconserved composition variable, coupled to a Newtonian stress relation for the reactant and product species. Multivalued reaction terms allow for different homogeneous phases to coexist with each other, resulting in banded composition and shear rate profiles. The one-dimensional equation of motion is evolved from a random initial state to its final steady state. We find that the system chooses banded states over homogeneous states, depending on the shape of the stress constitutive curve and the magnitude of the diffusion coefficient. Banding in the flow gradient direction under shear rate control is observed for shear-thinning transitions, while banding in the vorticity direction under stress control is observed for shear-thickening transitions. Received 1 April 2001 and Received in final form 16 June 2001