Reconstitution of the fusogenic activity of vesicular stomatitis virus.

Enveloped virus glycoproteins exhibit membrane fusion activity. We have analysed whether the G protein of vesicular stomatitis virus, reconstituted into liposomes, is able to fuse nucleated cells in a pH-dependent fashion. Proteoliposomes produced by octylglucoside dialysis did not exhibit cell fusion activity of the G protein. However, by making use of n-dodecyl octaethylene monoether (C12E8) as the solubilizing agent and by removal of the detergent in two steps, we were able to produce fusogenic G protein liposomes. These G protein liposomes fuse to the BHK-21 cell surface at pH 5.7-6.0 with an efficiency of fusion comparable with that of the parent virus. Physical and chemical analysis revealed that the fusogenic liposomes exhibited a protein to lipid weight ratio of 0.67 and showed an average diameter of 130 nm.     

Correlation between the order parameter and the steady-state fluorescence anisotropy of 1,6-diphenyl-1,3,5-hexatriene and an evaluation of membrane fluidity

In this paper it is shown that for 1,6-diphenyl-1,3,5-hexatriene there exists a simple analytical relation between the orientational order parameter and the steady-state fluorescence anisotropy. This relation is derived on semi-empirical grounds. The order parameter and the true microviscosity for membranes as calculated from steady-state measurements are evaluated. For biological membranes the estimation of the order parameter from steady-state experiments is feasible, but the evaluation of the true microviscosity is not reliable. Also, the physiological relevance of the order parameter is discussed.     

Double-tagged fluorescent bacterial bioreporter for the study of polycyclic aromatic hydrocarbon diffusion and bioavailability

Bacterial degradation of polycyclic aromatic hydrocarbons (PAHs), ubiquitous contaminants from oil and coal, is typically limited by poor accessibility of the contaminant to the bacteria. In order to measure PAH availability in complex systems, we designed a number of diffusion-based assays with a double-tagged bacterial reporter strain Burkholderia sartisoli RP037-mChe. The reporter strain is capable of mineralizing phenanthrene (PHE) and induces the expression of enhanced green fluorescent protein (eGFP) as a function of the PAH flux to the cell. At the same time, it produces a second autofluorescent protein (mCherry) in constitutive manner. Quantitative epifluorescence imaging was deployed in order to record reporter signals as a function of PAH availability. The reporter strain expressed eGFP proportionally to dosages of naphthalene or PHE in batch liquid cultures. To detect PAH diffusion from solid materials the reporter cells were embedded in 2 cm-sized agarose gel patches, and fluorescence was recorded over time for both markers as a function of distance to the PAH source. eGFP fluorescence gradients measured on known amounts of naphthalene or PHE served as calibration for quantifying PAH availability from contaminated soils. To detect reporter gene expression at even smaller diffusion distances, we mixed and immobilized cells with contaminated soils in an agarose gel. eGFP fluorescence measurements confirmed gel patch diffusion results that exposure to 2–3 mg lampblack soil gave four times higher expression than to material contaminated with 10 or 1 (mg PHE) g⁻¹.     

Lipid polarity and sorting in epithelial cells

Apical and basolateral membrane domains of epithelial cell plasma membranes possess unique lipid compositions. The tight junction, the structure separating the two domains, forms a diffusion barrier for membrane components and thereby prevents intermixing of the two sets of lipids. The barrier apparently resides in the outer, exoplasmic leaflet of the plasma membrane bilayer. First data are now available on the generation of these differences in Madin-Darby canine kidney (MDCK) cells, grown on filter supports. Experiments in which fluorescent precursors of apical lipids were introduced into the cell have demonstrated that upon biosynthesis apical lipids are sorted from basolateral lipids in an intracellular compartment. In this paper we present a model for the sorting process, the central point of which is that the two sets of lipids laterally segregate into microdomains that bud to form vesicles delivering the lipids to the apical and the basolateral plasma membrane domains, respectively.     

Biotechnological applications of Candida antarctica lipase A: State-of-the-art

The yeast Candida antarctica produces two different lipases, lipases A and B. While lipase B (CAL-B) is probably the mostly employed hydrolase in the biocatalysis field, the use of the lipase A (CAL-A) has been rather scarce and consequently its tridimensional structure has not been elucidated yet. However, CAL-A is a useful biocatalyst with many different applications that have been described especially in the last few years. Its attractiveness results from its unique features among hydrolases: the high thermostability, allowing operation at T > 90 °C; the ability to accept tertiary and sterically hindered alcohols, which has recently been attributed to the existence of a specific aminoacidic sequence in the active site; the sn-2 recognition in hydrolysis of triglycerides; the selectivity towards trans-fatty acids; the stability in the acidic pH range. Furthermore, it is considered to be an excellent biocatalyst for the asymmetric synthesis of amino acids/amino esters, due to its chemoselectivity towards amine groups. Considering all these aspects, in the present review, the origin, the properties and the applications of the CAL-A are briefly described and discussed, pointing out the unique characteristics of this biocatalyst.     

Looking up in Scotland? Multinationalism,multiculturalism and political elites

At a time when all the political parties of Scotland are trying to establish a persuasive vision of the nation, inquiry into where ethnic and racial minorities fit into these debates provides one understudied means of bridging literatures on multinationalism and multiculturalism. Focusing especially on the lesser known question of how elite political actors are positioning minorities within projects of nation-building, this article draws upon original empirical data in which three predominant clusters emerge. The first centres on an aspirational pluralism, in so far as political elites are less inclined – in contrast to counterparts in some other minority nations – to place ethnically determined barriers on membership of Scottish nationhood. The second concerns the competing ways in which the legacy of Scotland's place in the British Empire is appropriated by actors of different political hues, and so assumes a multiform role. The third cluster points to potential limitations in minority claims-making and recognition, especially in terms of formal multilingualism and corporate multifaithism, something that may partly be explained by the tension between multinationalism and multiculturalism. Taken together, the article illustrates how elite political actors can play a vital role in ensuring that appeals to nationhood in Scotland can be meaningfully calibrated to include minorities too.