酿酒酵母谷胱甘肽提取工艺条件的研究

研究了不同抽提方法对酿酒酵母中谷胱甘肽提取率的影响,确定微波法是提取谷胱甘肽的有效方法,微波法最优提取条件为：时间150S,火力为中高火,料液比1：160,优化后谷胱甘肽的提取率比以前提高了11．74％．并将提取的谷胱甘肽制成成品,采用X-射线衍射分析对其进行了检测．     

Enzymatic oxidation of alkoxyanilines for preparation of conducting polymers

A biomimetic route for synthesis of a conducting molecular complex between polyalkoxyanilines and a polyelectrolyte, poly (sodium 4‐styrenesulfonate) (SPS) is presented. Horseradish peroxidase (HRP) was used to catalyze the polymerization of alkoxyanilines. A few of water‐soluble ring‐substituted polyalkoxyanilines have been enzymatically synthesized with variation of groups, such as ortho‐methoxy, meta‐methoxy, ortho‐ethoxy, and meta‐ethoxy to form polyalkoxyanilines/SPS complexes. The presence of alkoxy substituents affects the polymerization reactions. These enzymatic oxidation reactions occur in a different potential range to that observed for the chemical polymerization. Similar electrochemical and optical properties were obtained for every pair of ortho‐ and meta‐alkoxy substituted polyanilines. For comparing, polyalkoxyanilines were also prepared by chemical polymerization in the presence of SPS. © 2006 Wiley Periodicals, Inc. J Appl Polym Sci 103: 103: 3724–3729, 2007     

Horseradish peroxidase‐catalysed oxidation of aqueous natural and synthetic oestrogens

The horseradish peroxidase (HRP)‐catalysed oxidation of selected potent oestrogens, including the natural oestrogens 17β‐oestradiol and oestriol and the synthetic oestrogen ethinyloestradiol, was studied and compared with that of phenol. HRP catalysed the oxidation of these oestrogens and phenol over a wide range of pH values, with optimal performance around neutral pH, which is in the range of typical urban wastewaters. In comparison with that of phenol, the oxidation of oestrogens consistently required less hydrogen peroxide. In addition, the rate of oxidation of oestradiol was equivalent, twofold faster and fivefold faster compared with that of ethinyloestradiol, oestriol and phenol respectively. For all substrates studied, similar kinetics of removal were observed provided that sufficient enzyme was added to reactions to compensate for differences in substrate affinity. In contrast with earlier studies with phenols in which HRP was observed to be susceptible to significant inactivation due to interactions with hydrogen peroxide and reaction products, minimal inactivation of HRP was observed during the present study, probably owing to the very low concentration of target substrates used here (i.e. 10 µmol dm^?3) relative to earlier studies with other phenolic substrates. These observations suggest that this enzymatic approach has strong potential to be used to target the treatment of oestrogenic compounds. Copyright © 2007 Society of Chemical Industry     

High production of glutathione by in vitro enzymatic cascade after thermostability enhancement

The cell free system has been paid more attention due to its potential of facilitating efficient catalysis of multistep reactions. In this study, an efficient enzymatic cascade of glutathione (GSH) production was developed through the evolution of bifunctional glutathione synthetase (GshF), coupled with polyphosphate kinase (PPK). First, the stability and activity of GshF were enhanced by loop interchange and site-directed mutagenesis. As a result, the GshF half-value period increased 163.32-fold, and its activity raised 18.16%. PPK from Jhaorihella thermophile (PPKJT) was characterized and used to regenerate ATP in the GSH synthesis, with hexametaphosphate (PolyP (6)) as the phosphate donor. After the process optimization, 99.92 mM GSH and 7.61 mM oxidized glutathione (GSSG) were produced within 2 hr. The molar yield was 0.96 mol/mol based on the amino acid added, while the productivities of GSH achieved 49.95 mM/hr, which were the highest yield and productivity ever reported about GSH synthesis.     

Hemizygous Granzyme A Mice Expressing the hSOD1G93A Transgene Show Slightly Extended Lifespan

Granzyme A (gzmA), a serine protease involved in the modulation of the inflammatory immune response, is found at an elevated level in the serum from ALS patients. However, the influence of gzmA on the progression of ALS remains unclear. The aim of our work was to assess whether the absence of gzmA in an ALS murine model could help slow down the progression of the disease. Homozygous and hemizygous gzmA-deficient mice expressing the hSOD1G93A transgene were generated, and survival of these mice was monitored. Subsequently, gene and protein expression of inflammatory and oxidative stress markers was measured in the spinal cord and quadriceps of these mice. We observed the longest lifespan in gzmA+/− mice. GzmA gene and protein expression was downregulated in the spinal cord and serum from gmzA+/− mice, confirming that the increased survival of hemizygous mice is correlated with lower levels of gzmA. In addition, mRNA and protein levels of glutathione reductase (GSR), involved in oxidative stress, were found downregulated in the spinal cord and quadriceps of gmzA+/− mice, together with lower IL-1β and IL-6 mRNA levels in hemyzigous mice. In summary, our findings indicate for the first time that reduced levels, but not the absence, of gzmA could slightly ameliorate the disease progression in this animal model.     

Observing How Glutathione and S-Hexyl Glutathione Bind to Glutathione S-Transferase from Rhipicephalus (Boophilus) microplus

Rhipicephalus (Boophilus) microplus is one of the most widespread ticks causing a massive loss to livestock production. The long-term use of acaracides rapidly develops acaracide resistance. In R. microplus, enhancing the metabolic activity of glutathione S-transferase (RmGST) is one of the mechanisms underlying acaracide resistance. RmGST catalyzes the conjugation of glutathione (GSH) to insecticides causing an easy-to-excrete conjugate. The active RmGST dimer contains two active sites (hydrophobic co-substrate binding site (H-site) and GSH binding site (G-site)) in each monomer. To preserve the insecticide efficacy, s-hexyl glutathione (GTX), a GST inhibitor, has been used as a synergist. To date, no molecular information on the RmGST-GSH/GTX complex is available. The insight is important for developing a novel RmGST inhibitor. Therefore, in this work, molecular dynamics simulations (MD) were performed to explore the binding of GTX and GSH to RmGST. GSH binds tighter and sits rigidly inside the G-site, while flexible GTX occupies both active sites. In GSH, the backbone mainly interacts with W8, R43, W46, K50, N59, L60, Q72, and S73, while its thiol group directs to Y7. In contrast, the aliphatic hexyl of GTX protrudes into the H-site and allows a flexible peptide core to form various interactions. Such high GTX flexibility and the protrusion of its hexyl moiety to the H-site suggest the dual role of GTX in preventing the conjugation reaction and the binding of acaracide. This insight can provide a better understanding of an important insecticide-resistance mechanism, which may in turn facilitate the development of novel approaches to tick control.