Reducible disulfide-based non-viral gene delivery systems

The design and synthesis of safe efficient non-viral vectors for gene delivery has attracted significant attention in recent years due primarily to the severe side-effect profile reported with the use of their viral counterparts. Previous experiments have revealed that the strong interaction between the carriers and nucleic acid may well hinder the release of the gene from the complex in the cytosol adversely affecting transfection efficiency. However, incorporating reducible disulfide bonds within the delivery systems themselves which are then cleaved in the glutathione-rich intracellular environment may help in solving this puzzle. This review focuses on recent development of these reducible carriers. The biological rationale and approaches to the synthesis of reducible vectors are discussed in detail. The in vitro and in vivo evaluations of reducible carriers are also summarized and it is evident that they offer a promising approach in non-viral gene delivery system design.

Selective in vitro glycosylation of recombinant proteins:semi-synthesis of novel homogeneous glycoforms of human erythropoietin

Background: A natural glycoprotein usually exists as a spectrum of glycosylated forms, where each protein molecule may be associated with an array of oligosaccharide structures. The overall range of glycoforms can have a variety of different biophysical and biochemical properties, although details of structure–function relationships are poorly understood, because of the microheterogeneity of biological samples. Hence, there is clearly a need for synthetic methods that give access to natural and unnatural homogeneously glycosylated proteins. The synthesis of novel glycoproteins through the selective reaction of glycosyl iodoacetamides with the thiol groups of cysteine residues, placed by site-directed mutagenesis at desired glycosylation sites has been developed. This provides a general method for the synthesis of homogeneously glycosylated proteins that carry saccharide side chains at natural or unnatural glycosylation sites. Here, we have shown that the approach can be applied to the glycoprotein hormone erythropoietin, an important therapeutic glycoprotein with three sites of N-glycosylation that are essential for in vivo biological activity. Results: Wild-type recombinant erythropoietin and three mutants in which glycosylation site asparagine residues had been changed to cysteines (His10-WThEPO, His10-Asn24Cys, His10-Asn38Cys, His10-Asn83CyshEPO) were overexpressed and purified in yields of 13 mg l−1 from Escherichia coli. Chemical glycosylation with glycosyl-β-N-iodoacetamides could be monitored by electrospray MS. Both in the wild-type and in the mutant proteins, the potential side reaction of the other four cysteine residues (all involved in disulfide bonds) were not observed. Yield of glycosylation was generally about 50% and purification of glycosylated protein from non-glycosylated protein was readily carried out using lectin affinity chromatography. Dynamic light scattering analysis of the purified glycoproteins suggested that the glycoforms produced were monomeric and folded identically to the wild-type protein. Conclusions: Erythropoietin expressed in E. coli bearing specific Asn→Cys mutations at natural glycosylation sites can be glycosylated using β-N-glycosyl iodoacetamides even in the presence of two disulfide bonds. The findings provide the basis for further elaboration of the glycan structures and development of this general methodology for the synthesis of semi-synthetic glycoproteins. Results: Wild-type recombinant erythropoietin and three mutants in which glycosylation site asparagine residues had been changed to cysteines (His10-WThEPO, His10-Asn24Cys, His10-Asn38Cys, His10-Asn83CyshEPO) were overexpressed and purified in yields of 13 mg l−1 from Escherichia coli. Chemical glycosylation with glycosyl-β-N-iodoacetamides could be monitored by electrospray MS. Both in the wild-type and in the mutant proteins, the potential side reaction of the other four cysteine residues (all involved in disulfide bonds) were not observed. Yield of glycosylation was generally about 50% and purification of glycosylated protein from non-glycosylated protein was readily carried out using lectin affinity chromatography. Dynamic light scattering analysis of the purified glycoproteins suggested that the glycoforms produced were monomeric and folded identically to the wild-type protein. Conclusions: Erythropoietin expressed in E. coli bearing specific Asn→Cys mutations at natural glycosylation sites can be glycosylated using β-N-glycosyl iodoacetamides even in the presence of two disulfide bonds. The findings provide the basis for further elaboration of the glycan structures and development of this general methodology for the synthesis of semi-synthetic glycoproteins

Estimating time-varying risk premia in UK long-term government bonds

Simple models of time-varying risk premia are used to measure the risk premia in long-term UK government bonds. The parameters of the models can be estimated using nonlinear seemingly unrelated regression (NL-SUR), which permits efficient use of information across the entire yield curve and facilitates the testing of various cross-sectional restrictions. The estimated time-varying premia are found to be substantially different to those estimated using models that assume constant risk premia. © 2004 Taylor and Francis Ltd.

Three-dimensional nitrogen-doped graphene supported molybdenum disulfide nanoparticles as an advanced catalyst for hydrogen evolution reaction

An efficient three-dimensional (3D) hybrid material of nitrogen-doped graphene sheets (N-RGO) supporting molybdenum disulfide (MoS2) nanoparticles with high-performance electrocatalytic activity for hydrogen evolution reaction (HER) is fabricated by using a facile hydrothermal route. Comprehensive microscopic and spectroscopic characterizations confirm the resulting hybrid material possesses a 3D crumpled few-layered graphene network structure decorated with MoS2 nanoparticles. Electrochemical characterization analysis reveals that the resulting hybrid material exhibits efficient electrocatalytic activity toward HER under acidic conditions with a low onset potential of 112 mV and a small Tafel slope of 44 mV per decade. The enhanced mechanism of electrocatalytic activity has been investigated in detail by controlling the elemental composition, electrical conductance and surface morphology of the 3D hybrid as well as Density Functional Theory (DFT) calculations. This demonstrates that the abundance of exposed active sulfur edge sites in the MoS2 and nitrogen active functional moieties in N-RGO are synergistically responsible for the catalytic activity, whilst the distinguished and coherent interface in MoS 2 /N-RGO facilitates the electron transfer during electrocatalysis. Our study gives insights into the physical/chemical mechanism of enhanced HER performance in MoS2/N-RGO hybrids and illustrates how to design and construct a 3D hybrid to maximize the catalytic efficiency.

Chemical reactions of double bonds in activated carbon:microwave and bromination methods

The reaction of localised C=C bonds on the surface of activated carbons has been shown to be an effective method of chemical modification especially using microwave-assisted reactions.

The rise and fall of diasporic bonds in Japanese-Peruvian “Return” migration

Two decades after Japanese-Peruvians and other South Americans of Japanese descent began to migrate back to Japan, the return-migration phenomenon has ended. Induced by the Japanese government in the name of shared ethnicity, Japanese policymakers now largely regard return migration as a failed policy. It failed because return-migrants did not, in the view of policy- makers, assimilate, integrate, or “make it” in Japan as expected. Thus, once-imagined ethnic bonds ceased to exist in Japan. However, ethnic bonds sustained themselves well outside Japan. The Japanese-Peruvian community in Peru has thrived and maintained continuous ties with Japan. What explains the rise and fall of diasporic ethnic bonds? Drawing on my ethnographic research in Japanese-Peruvian communities in Peru and Japan, I found that diasporic ethnic bonds are cultivated or weakened depending upon where diasporic populations are located in relation to their ancestral homeland, and how such ties are utilized, for what, and by whom.

Q-switched Raman fiber laser with molybdenum disulfide-based passive saturable absorber

We demonstrate a Q-switched Raman fiber laser using molybdenum disulfide (MoS2) as a saturable absorber (SA). The SA is assembled by depositing a mechanically exfoliated MoS2 onto a fiber ferrule facet before it is matched with another clean ferrule via a connector. It is inserted in a Raman fiber laser cavity with a total cavity length of about 8km to generate a Q-switching pulse train operating at 1560.2 nm. A 7.7-km-long dispersion compensating fiber with 584 ps·nm?1km?1 of dispersion is used as a nonlinear gain medium. As the pump power is increased from 395mW to 422mW, the repetition rate of the Q-switching pulses can be increased from 132.7 to 137.4 kHz while the pulse width is concurrently decreased from 3.35μs to 3.03 μs. The maximum pulse energy of 54.3 nJ is obtained at the maximum pump power of 422mW. These results show that the mechanically exfoliated MoS2 SA has a great potential to be used for pulse generation in Raman fiber laser systems.