Purification and Characterization of Thermostable Chitinase from <Emphasis Type="Italic">Bacillus licheniformis</Emphasis> SK-1

Chitinase was purified from the culture medium of Bacillus licheniformis SK-1 by colloidal chitin affinity adsorption followed by diethylamino ethanol-cellulose column chromatography. The purified enzyme showed a single band on sodium dodecyl sulfate polyacrylamide gel electrophoresis. The molecular size and pI of chitinase 72 (Chi72) were 72 kDa and 4.62 (Chi72) kDa, respectively. The purified chitinase revealed two activity optima at pH 6 and 8 when colloidal chitin was used as substrate. The enzyme exhibited activity in broad temperature range, from 40 to 70°C, with optimum at 55°C. It was stable for 2 h at temperatures below 60°C and stable over a broad pH range of 4.0–9.0 for 24 h. The apparent K _m and V _max of Chi72 for colloidal chitin were 0.23 mg ml⁻¹ and 7.03 U/mg, respectively. The chitinase activity was high on colloidal chitin, regenerated chitin, partially N-acetylated chitin, and chitosan. N-bromosuccinamide completely inhibited the enzyme activity. This enzyme should be a good candidate for applications in the recycling of chitin waste.     

Acene-doped polymer films: singlet oxygen dosimetry and protein sensing

This paper describes thin films comprising acenes dispersed in a conjugated polymeric host that have a ratiometric photoluminescence response to singlet oxygen. These films also respond to irradiation of protein-bound sensitizers, which represents a solution to the problem of protein-conjugated polymer non-specific interactions.     

Gaussian modification of neutrino energy loss on strongly screening nuclides ~（55）Co and ~（56）Ni by electron capture in stellar interior

Gaussian modifications of the neutrino energy loss (NEL) by electron capture on the strongly screening nuclides ⁵⁵Co and ⁵⁶Ni are investigated. The results show that in strong electron screening (SES), the NEL rates decrease without modifying the Gamow-Teller (G-T) resonance transition. For instance, the NEL rates of ⁵⁵Co and ⁵⁶Ni decrease more than two and three orders of magnitude for ρ₇=5.86, T₉≤5, Y_e=0.47, Δ=6.3, respectively. In contrast, due to Gaussian modification, the NEL rates increase about two orders of magnitude in SES. Due to SES, the maximum values of the C-factor (in %) on NEL of ⁵⁵Co, ⁵⁶Ni are of the order of 99.80%, 99.56% at ρ₇=5.86 Y_e=0.47 and 99.60%, 99.65% at ρ₇=106 Y_e=0.43, respectively.     

Microbubble mediated sonoporation of cells in suspension: Clonogenic viability and influence of molecular size on uptake

This work investigates whether the application of sonoporation is limited by the size of a macromolecule being delivered and by the ability of cells to proliferate following uptake. KHT-C cells in suspension were exposed to variations in ultrasound pressure (0-570 kPa) and microbubble shell-type (lipid and protein) at fixed settings of 500 kHz centre frequency, 32 μs pulse duration, 3 kHz pulse repetition frequency and 2 min insonation. Reversible permeability (P_R), defined as the number of cells stained with FITC-dextran and unstained with propidium iodide (i.e., PI-viable), was measured with flow cytometry for marker molecules ranging from 10 kDa to 2 MDa in size. Viable permeability (P_V) defined as the number of permeabilised cells that maintained their ability to proliferate, was measured by clonogenic assay. Comparable intracellular delivery of all sizes of molecules was achieved, indicating that intracellular delivery of common therapeutic drugs may not be limited by molecular size. Maximum P_R’s of 80% (at 10 kDa) and 55% (at 10 kDa) were achieved with lipid coated bubbles at 3.3% v/v and protein coated bubbles at 6.7% v/v concentrations. The PI-viability was approximately 80% at 570 kPa in both cases. The maximum P_V achieved with both agents was 22%, while inducing a lower overall clonogenic viability with the lipid (39%) compared to the protein (56%) shelled bubbles. This study demonstrates that large macromolecules, up to 2 MDa in size, can be delivered with high efficiency to cells which undergo reversible permeabilisation, maintaining long-term viability in approximately half of the cells.     

循环矩阵环的局部化

本文的主要结果如下：(1)环R关于其乘法封闭子集S满足左Ore条件当且仅当R[σ1,σ2,…，σt]关于其相应乘法封闭子集S[σ1,σ2,…，σt]满足左Ore条件．(2)若R关于其乘法封闭子集S满足左Ore条件，S^-1 R是R关于S的左分式环，其自然同态为φ：R→S^-1R，则存在环同态φ：R[σ1,σ2,…,σt]→S[σ1,σ2,…,σt]^-1 R[σ1,σ2,…σt]使得(S-1R)[φ（σ1),φ-(σ2),…φ（σt)]≌S[σl，σ2，…，σt]^-1R[σ1,σ2,…σt]。     

Triply fuzzy function approximation for hierarchical Bayesian inference

We prove that three independent fuzzy systems can uniformly approximate Bayesian posterior probability density functions by approximating the prior and likelihood probability densities as well as the hyperprior probability densities that underly the priors. This triply fuzzy function approximation extends the recent theorem for uniformly approximating the posterior density by approximating just the prior and likelihood densities. This approximation allows users to state priors and hyper-priors in words or rules as well as to adapt them from sample data. A fuzzy system with just two rules can exactly represent common closed-form probability densities so long as they are bounded. The function approximators can also be neural networks or any other type of uniform function approximator. Iterative fuzzy Bayesian inference can lead to rule explosion. We prove that conjugacy in the if-part set functions for prior, hyperprior, and likelihood fuzzy approximators reduces rule explosion. We also prove that a type of semi-conjugacy of if-part set functions for those fuzzy approximators results in fewer parameters in the fuzzy posterior approximator.     

Expression of UDP-glucose dehydrogenase reduces cell-wall polysaccharide concentration and increases xylose content in alfalfa stems

The primary cell-wall matrix of most higher plants is composed of large amounts of uronic acids, primarily d-galacturonic acid residues in the back-bone of pectic polysaccharides. Uridine diphosphate (UDP)-glucose dehydrogenase is a key enzyme in the biosynthesis of uronic acids. We produced transgenic alfalfa (Medicago sativa) plants expressing a soybean UDP-glucose dehydrogenase cDNA under the control of two promoters active in alfalfa vascular tissues. In initial greenhouse experiments, enzyme activity in transgenic lines was up to seven-fold greater than in nontransformed control plants; however, field-grown transgenic plants had only a maximum of 1.9-fold more activity than the control. Cell-wall polysaccharide content was lower and Klason lignin content was higher in transgenics compared to the nontransformed control. No significant increase in pectin or uronic acids in the polysaccharide fraction was observed in any line. Xylose increased 15% in most transgenic lines and mannose concentration decreased slightly in all lines. Because of the complexity of pectic polysaccharides and sugar biosynthesis, it may be necessary to manipulate multiple steps in carbohydrate metabolism to alter the pectin content of alfalfa.     

Combinatorial library of peptidotriazoles: identification of [1,2,3]-triazole inhibitors against a recombinant Leishmania mexicana cysteine protease

A library consisting of about half of 800 000 possible peptidotriazoles on 450 000 beads was prepared by solid-phase peptide synthesis combined with a regiospecific copper(I)-catalyzed 1,3-dipolar cycloaddition between a resin-bound alkyne and a protected amino azide. The central [1,2,3]-triazole was flanked on each side by two randomized amino acids introduced in a combinatorial approach. Importantly, the formation of the triazole could be performed quantitatively in a randomized fashion. The library was screened on solid phase for inhibitory effect against a recombinant cysteine protease, Leishmania mexicana CPB2.8DeltaCTE and sorted by a high-throughput instrument, COPAS beadsorter (up to 200 000 beads/h). Forty-eight hits were analyzed by MALDI-TOF MS providing structural information about the protease specificity, and 23 peptidotriazoles were resynthesized and evaluated in solution, with the best inhibitor displaying a K(i) value of 76 nM. A one-pot procedure was used to convert Fmoc-amino azides into their corresponding Boc derivatives. The crucial influence of weak interactions with a spacer used for detection by MALDI-TOF MS on screening results was observed.