A chimeric gene, Glu-Xyl, encoding Bacillus amyloliquefaciens glucanase (Glu, 24.4 kDa) and Bacillus subtilis xylanase (Xyl, 21.2 kDa), was constructed via end-to-end fusion and expressed successfully in Escherichia coli. The purified fusion protein (46.1 kDa) exhibited both glucanase and xylanase activities. Compared with parental enzymes, the Glu moiety was characterized by kinetic parameters of decreased K(m) (0.66-fold) and increased K(cat) (2.75-fold), whereas the Xyl moiety had an increased K(m) (1.37-fold) and decreased K(cat) (0.79-fold). These indicate a 3.15-fold net increase and a 31% decrease in catalytic efficiency (K(cat)/K(m)) of the Glu and Xyl moieties. Activities and stabilities of both moieties at 40-90 degrees C or pH 3.0-10.0 were compared with those of the parental enzymes. Despite some variations, common optima were 40 degrees C and pH 9.0 for the Glu moiety and parent, and 50-60 degrees C and pH 9.0 for the Xyl counterparts. Thus, the fusion enzyme Glu-Xyl was bifunctional, with greatly enhanced glucanase activity associated with a decrease in xylanase activity.

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We investigate the transport diffusion of methane at 300 K and pressures of up to 15 bar in dry and wetted carbon nanotubes (CNTs) having diameters ranging from 0.95 to 2.034 nm using nonequilibrium molecular dynamics (NEMD) simulation. Because of their strong hydrogen bonding, preadsorbed water molecules transport in the form of clusters and block the diffusion of methane, reducing the Onsager coefficient of methane dramatically compared to that in dry CNTs. The reduction in the methane Onsager coefficient is greater in narrower CNTs or at higher water densities. Because the diameter of the water clusters is almost invariant with water density, the Onsager coefficient of water in the (10, 10) CNT increases linearly with water density. It is further found that whereas decreasing the CNT diameter from 2.034 to 0.95 nm enhances the Onsager coefficient of pure methane by about 1 order of magnitude, the Onsager coefficient of water is almost independent of the CNT diameter at a water density of 0.05 g/cm. We propose a theoretical model for the strong dependency of methane diffusion in wetted CNTs on the Onsager coefficient of water, the preadsorbed water density, and the CNT diameter. The model predicts the Onsager coefficients of the methane/water mixture from the Onsager coefficients of the pure components. Our study provides a basic understanding of the coupled diffusion of immiscible components in nanochannels and will facilitate progress in gas storage and carbon capture as well as nanofiltration and biomedical and biotechnological applications.

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