Protein nanotubes with an enzyme interior surface

This report describes the synthesis and enzyme activities of multilayered protein nanotubes with an α-glucosidase (αGluD) interior surface. The nanotubes were prepared by using an alternating layer-by-layer (LbL) assembly of human serum albumin (HSA) and oppositely charged poly-L-arginine (PLA) into a track-etched polycarbonate (PC) membrane (pore size=400 nm) followed by addition of αGluD as the last layer of the wall. Subsequent dissolution of the PC template yielded (PLA/HSA)(2)PLA/αGluD nanotubes. SEM measurements revealed the formation of uniform hollow cylinders with (413±17) nm outer diameter and (52±3) nm wall thickness. In aqueous media, the nanotubes captured a fluorogenic glucopyranoside, 4-methyl-umbelliferyl-α-D-glucopyranoside (MUGlc), into their one-dimensional pore space and hydrolyzed the substrate efficiently to form α-D-glucose. We determined the enzyme parameters (Michaelis constant, K(M), and catalytic constant, k(cat), values) of the protein nanotubes. The several-micrometers-long cylinders were of sufficient length to be spun down by centrifugation at 4000 g, so the product could therefore be easily separated. Similar biocatalysts were prepared by complexation of biotinylated-αGluD into HSA-based nanotubes bearing a single avidin layer as an internal surface. The obtained hybrid nanotubes also exhibited the same enzyme activity for the MUGlc hydrolysis.     

Development of a temperature-variable magnetic resonance imaging system using a 1.0T yokeless permanent magnet

A temperature variable magnetic resonance imaging (MRI) system has been developed using a 1.0 T permanent magnet. A permanent magnet, gradient coils, radiofrequency coil, and shim coil were installed in a temperature variable thermostatic bath. First, the variation in the magnetic field inhomogeneity with temperature was measured. The inhomogeneity has a specific spatial symmetry, which scales linearly with temperature, and a single-channel shim coil was designed to compensate for the inhomogeneity. The inhomogeneity was drastically reduced by shimming over a wide range of temperature from −5 °C to 45 °C. MR images of an okra pod acquired at different temperatures demonstrated the high potential of the system for visualizing thermally sensitive properties.     

Synthesis of smooth and superconducting (Cu, C)–Ba–O/CaCuO2/(Cu, C)–Ba–O films using SrCuO2 buffer

Artificially stacked structures of [(infinite layer CaCuO₂)_k/(superconducting (Cu, C)Ba₂CuO_x:(Cu, C)-1201)_l]_m were fabricated on SrCuO₂ buffer layer by means of sequential deposition of each block using pulsed laser deposition. Smooth and epitaxial growth all over the deposition cycles of (Cu, C)-1201 and CaCuO₂ were confirmed by streak patterns of in situ RHEED. In comparison with (Cu, C)-1201 single layer films, the artificial stacking resulted in a rise of superconducting properties, T_c-onset and T_c(ρ=0) up to 95 K and 65 K, respectively.     

New Stereoselective GC Phases: Immobilized Chiral Polysiloxanes with (S)-(–)-t-Leucine Derivatives as Selectors

New chiral polysiloxanes have been prepared as stationary phases for gas chromatography, with (S)-(–)-t-leucine-t-butylamide, (S)-(–)-t-leucine-(S)-(–)-1-phenylethylamide, (S)-(–)-t-leucine-(S)-(–)-1-(α-naphthyl)ethylamide, (S)-(–)-t-leucine-(R)-( + )-1-phenylethylamide, and (S)-(–)-t-leucine-(R)-( + )-1-(α-naphthyl)ethylamide as selectors. Immobilization is achieved by radical-induced cross-linking with 1,3,5,7-tetravinyl-1,3,5,7-tetramethylcyclotetrasiloxane (V₄) and dicumyl peroxide (DCUP) as cross-linking reagents and cured at 170°C. Under these conditions, racemization of (S)-(–)-t-leucine is less than 4.5% (R) for 1 h curing, while for polysiloxanes with the conventional (S)-(–)-valine selectors about 20% of R-enantiomers are formed by racemization. In the presence of 5% (w/w) V₄ and 6% of DCUP with regard to the phases, 70–80% immobilization is achieved; without V₄, the degree of immobilization is about 50% for both the (S)-(–)-t-leucine and (S)-(–)-valine selectors. As the size of the amide moieties of the selectors increases from t-butyl to 1-(α-naphthyl)ethyl, the degree of immobilization decreases. If the curing time is prolonged to 2 h, the extent of racemization increases. The selectivity factors achieved for amino acid enantiomers and similar pharmaceuticals are generally higher than those obtained with the corresponding non-immobilized Chirasil-Val phases.     

The Methionine 549 and Leucine 552 Residues of Friedelin Synthase from Maytenus ilicifolia Are Important for Substrate Binding Specificity

Friedelin, a pentacyclic triterpene found in the leaves of the Celastraceae species, demonstrates numerous biological activities and is a precursor of quinonemethide triterpenes, which are promising antitumoral agents. Friedelin is biosynthesized from the cyclization of 2,3-oxidosqualene, involving a series of rearrangements to form a ketone by deprotonation of the hydroxylated intermediate, without the aid of an oxidoreductase enzyme. Mutagenesis studies among oxidosqualene cyclases (OSCs) have demonstrated the influence of amino acid residues on rearrangements during substrate cyclization: loss of catalytic activity, stabilization, rearrangement control or specificity changing. In the present study, friedelin synthase from Maytenus ilicifolia (Celastraceae) was expressed heterologously in Saccharomyces cerevisiae. Site-directed mutagenesis studies were performed by replacing phenylalanine with tryptophan at position 473 (Phe473Trp), methionine with serine at position 549 (Met549Ser) and leucine with phenylalanine at position 552 (Leu552Phe). Mutation Phe473Trp led to a total loss of function; mutants Met549Ser and Leu552Phe interfered with the enzyme specificity leading to enhanced friedelin production, in addition to α-amyrin and β-amyrin. Hence, these data showed that methionine 549 and leucine 552 are important residues for the function of this synthase.     

Thermodynamic interactions in rubbery polymer/gas systems

The Flory–Huggins interaction parameters χ for 23 gases (He, Ne, Ar, Kr, Xe, H₂, N₂, O₂, N₂O, CO₂, CH₄, C₂H₄, C₂H₆, C₃H₆, C₃H₈, 1,3-C₄H₆, four C₄H₈'s, n-C₄H₁₀, iso-C₄H₁₀, and n-C₅H₁₂) in five rubbery polymers (1,2-polybutadiene (PB), poly(ethylene-co-vinyl acetate)) (EVAc), polyethylene (PE), polypropylene (PP), and poly(dimethyl siloxane) (PDMS) were determined from either literature data on Henry's law coefficient and partial molar volume or those on sorptive dilation for each polymer/gas system. Values of χ for the gases increased in the order of PDMS < PP ≡ PB < EVAc ≡ PE. Among the gases except He and H₂ whose χ values are not reliable, Ne and Xe have respectively the highest and the lowest values of χ for the polyolefins. The χ values of the hydrocarbons were compared together with previously reported χ values of n-alkanes C₃-C₁₀. The dependencies of χ upon concentration and temperature were discussed on the basis of the literature data. © 1997 John Wiley & Sons, Inc. J Polym Sci B: Polym Phys 35 : 1049–1053, 1997     

Formation of interpenetrated spherulites based on miscible crystalline polymer blends

The morphology and formation process of interpenetrated spherulites of poly(butylene succinate)/poly(vinylidene choloride‐co‐vinyl chloride) (PBSU/PVDCVC) blends were investigated by confocal laser scanning microscopy (CLSM). CLSM images showed that the dense fibrils of PBSU spherulites penetrated into the sparse PVDCVC spherulites. For a blend with PBSU content 50% and crystallization temperature T_c = 368 K, the simultaneous growth of PBSU and PVDCVC spherulites was observed. After PBSU fibrils collided with PVDCVC spherulites, they kept growing through PVDCVC spherulites. For a blend with PBSU content 30% and T_c = 363 K, PBSU started to nucleate after PVDCVC spherulites filled the whole space.