Dye-Affinity Nanofibrous Membrane for Adsorption of Lysozyme: Preparation and Performance Evaluation.

Polyacrylonitrile (PAN) nanofibrous membrane was prepared by an electrospinning technique. After heat treatment and alkaline hydrolysis, the weak ion exchange membrane was grafted with chitosan molecule and then covalently immobilized with a Cibacron Blue F3GA (CB). Fibre diameter, porosity and pore size of the membrane and immobilized dye density were characterized. Furthermore, the membrane was applied to evaluate the binding performance of lysozyme under various operating parameters (pH, chitosan mass per volume ratio, dye concentration, ionic strength and temperature) in batch mode. The experimental results were directly applied to purify lysozyme from chicken egg white by membrane chromatography. The results showed that the capture efficiency, recovery yield and purification factor were 90 and 87%, and 47-fold, respectively, in a single step. The binding capacity remained consistent after five repeated cycles of adsorption-desorption operations. This work demonstrates that the dye-affinity nanofibrous membrane holds great potential for purification of lysozyme from real feedstock.

ß-Chitin nanofibrils for self-sustaining hydrogels preparation via hydrothermal treatment.

A transparent nanofibril suspension could be readily obtained by treating purified squid pen powder in water with ultrasonic irradiation. The obtained suspension is consisted of ß-chitin nanofibrils (CNF) with 3-10 nm in width and several micrometers in length. The degree of acetylation (DA) of CNF was found to be 84% which is about 10% lower than that of untreated sample. The CNF suspension could be transformed into a durable 3-D hydrogels (CH) by simply heating to 180 °C for 1-4 h in an autoclave. Hydrophobic interaction between CNF was believed to play the major role for CNF self-assembling into hydrogels, since the as-prepared chitin hydrogels readily dissolved in a typical chaotropic solution (8 M urea) under room temperature. The hydrothermal duration and CNF concentration (0.3-2% (w/v)) strongly affected the physical properties of CH. The suspension of 1% (w/v) CNF treated with 4 h, 180 °C hydrothermal heating generated a CH with 99.3% water content, CNF with 87% crystallinity and an mechanical strength of 0.7 N breaking force.

Investigating the effects of sodium dodecyl sulfate on the aggregative behavior of hen egg-white lysozyme at acidic pH.

The research presented here is aimed at examining the effects of sodium dodecyl sulfate on the aggregative behavior of hen egg-white lysozyme at pH 2.0. Through various spectroscopic techniques, dynamic light scattering, and electron microscopy, we first demonstrated that SDS exhibited a biphasic effect on lysozyme fibrillation. The presence of SDS at higher concentrations (e.g., 0.25, 5.00, or 20.00 mM SDS) was found to suppress fibril formation of lysozyme whereas fibrillogenic lysozyme-SDS ensemble containing beta-sheet-rich conformation was observed upon the addition of lower concentrations of SDS (e.g., 0.00, 0.06, or 0.1mM SDS). Next, our equilibrium urea-unfolding data revealed that lysozyme samples with higher SDS concentrations showed superior thermodynamic stabilities over the ones with no or lower levels of SDS. Finally, the correlation between SDS concentration and lysozyme aggregative/fibrillogenic propensity and the underlying interacting mechanism were further explored using surface tensiometry and isothermal titration calorimetry. We believe the outcome from this work may not only help decipher the molecular mechanism of amyloid fibrillation, but also shed light on a rational design of potential therapeutic strategies for amyloid pathology.

Efficient and stable enzyme immobilization in a block copolypeptide vesicle-templated biomimetic silica support.

We report the immobilization of a model enzyme, papain, within silica matrices by combining vesiclization of poly-l-lysine-b-polyglycine block copolypeptides with following silica mineralization. Our novel strategy utilizes block polypeptide vesicles to induce the condensation of orthosilicic acid while trapping an enzyme within and between vesicles. The polypeptide mediated silica-immobilized enzyme exhibits enhanced pH and thermal stability and reusability, comparing with the free and vesicle encapsulated enzyme. The enhanced enzymatic activity in the immobilized enzyme is due to the confinement of the enzyme in the polypeptide mediated silica matrices. Kinetic analysis shows that the enzyme functionality is determined by the structure and property of silica/polypeptide matrices. The proposed novel strategy provides an alternative route for the synthesis of a broad range of functional bionanocomposites entrapped within silica nanostructures.

The epigenetic effects of amyloid-beta(1-40) on global DNA and neprilysin genes in murine cerebral endothelial cells.

Amyloid-beta (Abeta) is the core component of senile plaques, which are the pathological markers for Alzheimer's disease and cerebral amyloid angiopathy. DNA methylation/demethylation plays a crucial role in gene regulation and could also be responsible for presentation of senescence. Oxidative stress, which may be induced by Abeta, is thought to be an important contributor of DNA hyper-methylation; however, contradicting this is the fact that global DNA hypo-methylation has been found in aging brains. It therefore remains largely unknown as to whether Abeta does in fact cause DNA methylation/demethylation. Neprilysin (NEP) is one of the enzymes responsible for Abeta degradation, with its expression decreasing in both Alzheimer and aging brains. Using high-performance liquid chromatography (HPLC), we explore whether Abeta is responsible for alteration of the global DNA methylation status on a murine cerebral endothelial cells model, and also use methylation-specific PCR (MSPCR) to examine whether DNA methylation status is altered on the NEP promoter region. We find that Abeta reduces global DNA methylation whilst increasing NEP DNA methylation and further suppressing the NEP expression in mRNA and protein levels. Our results support that Abeta induces epigenetic effects, implying that DNA methylation may be part of a vicious cycle involving the reduction in NEP expression along with a resultant increase in Abeta accumulation, and that Abeta may induce global DNA hypo-methylation.