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Artificial planar bilayer lipid membranes (BLMs) are simple models of cellular systems under physically and chemically controlled conditions, and they have been used to investigate membrane protein activity. Baculovirus-budded virus (BV) systems can express recombinant membrane proteins. In this study, aiming for membrane protein reconstitution, we examined the fusion of BVs containing recombinant membrane proteins into artificial planar BLMs on a Si microwell substrate. BV fusion with the BLMs depended on the pH of the solution, and it was enhanced at lower pH. Based on fluorescence recovery after photobleaching (FRAP) measurement, the fusion state of BVs was evaluated, and full fusion at low pH was confirmed. The fluorescent labeling the membrane proteins was also observed in the freestanding part of the BLMs as well as in the supported part. These results demonstrate the effectiveness of BLMs as a platform to examine detailed fusion dynamics of BVs. Furthermore, this study revealed that the fusion of BVs is a promising method for reconstituting membrane proteins to artificial freestanding BLMs for the development of biodevices with which we can examine membrane protein activity.

The lateral diffusion of lipids is a key factor when functionalizing artificial planar bilayer lipid membranes (BLMs). Fluorescence recovery after photobleaching (FRAP) is an established method for evaluating the fluidity of BLMs by the quantitative determination of the diffusion coefficient. However, a BLM with a uniform diffusion coefficient is usually assumed for analysis. In addition, when the BLM to be evaluated is small, the spread of a bleached circle caused by lateral diffusion during the bleaching process and the divergence of the laser used for bleaching interfere with the quantitative analysis. In this study, a numerical calculation was adopted to make it possible to analyze the continuous BLM between freestanding and supported areas, where the diffusion coefficients change depending on the presence or absence of an interaction with the substrate. A quantitative evaluation independent of such bleaching conditions as the bleaching diameter was also ensured by incorporating the spreading effect of the bleached circle in the calculation, which was employed to analyze a freestanding BLM with a diameter of only a few micrometers. By comparing calculations and experiments on FRAP recovery curves, we found that there are multiple diffusion elements and high diffusion barriers at the boundary between the freestanding and supported areas in a BLM over a SiO 2 /Si microwell substrate.

The localization of liquid-ordered (L o ) and liquid-crystalline (L α ) phase domains on a silicon substrate with a microwell array is investigated. Although the phase separation of the L o and L α phases on both a giant unilamellar vesicle (GUV) and a supported membrane remains stable for a long time, the lateral diffusion of lipids across each domain boundary occurs quickly. Since the phase separation and domain arrangement are governed by the stiffness and lateral tension of the lipid membrane, the phase separation is rearranged on a micropatterned substrate. Similar phase separation of the L o and L α phases is observed at a lipid membrane suspended over a microwell. However, the L α phase is preferred at a suspended membrane, and saturated lipids and cholesterol are excluded toward the supporting membrane on the periphery. Since the L o domain area is reduced by anisotropic diffusion through the boundary between the suspended and supported membranes, a very slow reduction rate with a linear functional relation is observed. Finally, a localized L α phase domain is observed at a membrane suspended over a microwell, which is surrounded by an L o phase supported membrane.

Gold nanoparticles are generally considered to be biologically inactive. However, in this study we show that the addition of 1.4 nm diameter gold nanoparticle induces the remodeling of the ring-shaped protein TRAP into a hollow, capsid-like configuration. This structural remodeling is dependent upon the presence of cysteine residues on the TRAP surface as well as the specific type of gold nanoparticle. The results reveal an apparent novel catalytic role of gold nanoparticles., (© 2012 American Chemical Society)

We fabricated a heterogeneous supported lipid bilayer (SLB) by employing binary lipid mixtures comprising a saturated acyl chain DSPC and an unsaturated acyl chain nickel-chelating lipid. By using the specific adsorption properties of histidine-tagged proteins (His-tagged GFPs) in relation to nickel-chelating lipids, we demonstrated protein pattern formation on the SLB corresponding to the phase separation pattern of the SLB. In addition, by using a lipid mixture consisting of an unsaturated acyl chain DOPC and a nickel-chelating lipid, and His-tagged GFPs, we succeeded in transporting the proteins along a hydrophilic micropattern on a SiO(2) substrate. The protein transport is induced by the self-spreading behavior of a fluid SLB with a kinetic spreading coefficient beta = 10.4 microm(2) s(-1). This method provides a guide for strategically carrying various biomolecules to specific positions by using a soft biointerface on a solid surface. In addition, the results demonstrate the importance of using techniques that allow the controlled manipulation of biomolecules based on the static or dynamic properties of the SLB platform.

We report on the self-spreading behavior of a supported lipid bilayer (SLB) on a silicon surface with various 100 nm nanostructures. SLBs have been successfully grown from a small spot of a lipid molecule source both on a flat surface and uneven surfaces with 100 nm up-and-down nanostructures. After an hour, the self-spreading SLB forms a large circle or an ellipse depending on the nanostructure pattern. The results are explained by a model that shows that a single-layer SLB grows along the nanostructured surfaces. The model is further supported by a quantitative analyses of our data. We also discuss the stability of the SLB on nanostructured surfaces in terms of the balance between its bending and adhesion energies.

The new europium(III) chelate [2,2',2'',2'''-[[4'-(aminobiphenyl-4-yl)-2,2':6',2''-terpyridine- 6,6''-diyl]bis(methylenenitrilo)]tetrakis(acetato)] europium(III) (ATBTA-Eu3+) and its 4,6-dichloro-1,3,5-triazinyl and succinimidyl derivatives (DTBTA and NHS-ATBTA, respectively) were synthesized and characterized. Both labeling complexes DTBTA-Eu3+ and NHS-ATBTA-Eu3+ are luminescent. Especially DTBTA-Eu3+ is strongly luminescent, with a luminescence quantum yield of 9.1%, molar extinction coefficient of 3.1 x 10(4) cm(-1) M(-1) (335 nm), and luminescence lifetime of 1.02 ms. The excitation and emission maximum wavelengths of DTBTA-Eu3+ are 335 and 616 nm, respectively. The complex is very stable in aqueous buffers, with a conditional formation constant log K(DTBTA-Eu) of 25.0 at pH 8, and can be conjugated to DNA and proteins. The chelates are also highly resistant to thermal decomposition, photodegradation, and ozone oxidation. These properties prove that DTBTA-Eu3+ is suitable as a luminescence label in DNA assays.

[reaction: see text] A novel multiregulation system of ion recognition was constructed by utilizing a molecular gate. Interaction between the host and an Ag(+) is controlled with an Na(+) ion.