Altered expression of polyhydroxyalkanoate synthase gene and its effect on poly[(R)-3-hydroxybutyrate] synthesis in recombinant Escherichia coli

Polyhydroxyalkanoate (PHA) synthase (PhaC) from Wautersia eutropha was expressed in a wide range of production level in Escherichia coli XL1-Blue cells and its effects on PhaC activity, poly[(R)-3-hydroxybutyrate] [P(3HB)] production and its molecular weights were investigated. The production level of PhaC was controlled both by the amount of chemical inducer (isopropyl-β-d-thiogalactopyranoside, IPTG) added into the medium and the use of different copy number of plasmids. In a flask experiment, as PhaC production level in the cells increased, the PhaC activity also increased in the range of low PhaC concentration. However, PhaC activity did not further increase in the range of high PhaC concentration, probably due to the formation of inclusion body in the cells. The molecular weight of P(3HB) was found to decrease with increasing PhaC activity. This trend was also verified in high cell density cultivation using 10-l jar fermentor. Furthermore, we demonstrated that the use of low copy number plasmid and appropriate induction of PhaC expression were effective in achieving both high productivity and high molecular weight of P(3HB).     

Prostaglandin D2 synthase and its post-translational modifications in neurological disorders

Prostaglandin D2 synthase (PGDS) (beta-trace protein) is a highly abundant cerebrospinal fluid (CSF) glycoprotein. A number of studies have been performed to determine the potential value of this protein for the diagnosis of various neurological disorders. The measurement of total PGDS levels in CSF has proved marginally useful for this purpose, but promising results were obtained while investigating changes in the posttranslational modifications (PTM) pattern. Using 2-DE analysis, we previously showed that PGDS is differentially expressed in ante- and post mortem CSF samples. In the present study, we examined whether the PGDS isoforms may help to distinguish stroke and neurodegenerative disease patients from healthy subjects. The pattern of PGDS PTM was analyzed in CSF from patients with various neurological disorders (n = 44) using IEF/immunoblotting techniques. Strong alterations of this pattern were detected in patients with different forms of degenerative dementia. These findings are consistent with PGDS being altered in some neurological diseases and provide new opportunities for clinical applications.     

Photochemical reactions of a dimethacrylate compound containing a chalcone moiety in the main chain

We synthesized the new photosensitive oligomer containing a chalcone moiety in the main chain by end-capping reaction of diepoxide compound with methacrylic acid. The chalcone-epoxy oligomeric compound was synthesized with 4,4^′-dihydroxychalcone and epichlorohydrin. Investigation of the photosensitivity of the newly synthesized chalcone oligomer was carried out by using UV-Vis absorption and infrared spectroscopies under UV exposure. We observed the photodimerization behavior under UV irradiation. At the same time, we could also observe the photopolymerization of the compound with a trace amount of dimethoxyphenyl acetophenone. Thermal properties of UV-cured dimethacrylate compounds were also studied.     

Genome-inspired search for new antibiotics. Isolation and structure determination of new 28-membered polyketide macrolactones, halstoctacosanolides A and B, from Streptomyces halstedii HC34

During the search for polyketide synthase (PKS) in the genome of Streptomyces halstedii HC34, we found clustered new genes which appeared to encode typical Type 1 PKSs beyond the cluster harboring the genes for the biosynthesis of antitumor antibiotic vicenistatin. The deduced domain configuration of these putative PKS genes allowed to predict a corresponding partial structure of polyketide, which was in turn materialized by isolation of new polyketide macrolactone halstoctacosanolides A and B from the fermentation broth of S. halstedii HC34. The structures of these metabolites were determined by spectroscopic means to have a novel 28-membered macrolactone structure. The partial structure deduced from the genetic data was completely compatible to the structures of halstoctacosanolides A and B. This success clearly demonstrates the present new approach of genome-inspired search for new antibiotics promising. Halstoctacosanolides A and B showed moderate antimicrobial activity against several microorganisms.     

含氧膦结构的可溶性感光聚酰亚胺合成与表征

聚酰亚胺(PI)是一类综合性能优异的功能性材料,广泛应用于航天、航空及电子工业等领域[1].感光聚酰亚胺可以采用光刻工艺,大大简化了其应用加工程序[2,3],因而备受人们所青睐.含查尔酮结构的聚合物对UV辐射敏感度高和化学稳定性好,最近被广泛的研究和应用[4～7].主链含查尔酮结     

Construction of recombinant Bacillus subtilis for production of polyhydroxyalkanoates

Polyhydroxyalkanoates (PHAs) are polyesters of hydroxyalkanoates synthesized by numerous bacteria as intracellular carbon and energy storage compounds and accumulated as granules in the cytoplasm of cells. In this work, we constructed two recombinant plasmids, pBE2C1, and pBE2C1AB, containing one or two PHA synthse, genes, respectively. The two plasmids were inserted into Bacillus subtilis DB104 to generate modified strains, B. subtilis/pBE2C1 and B. subtilis/pBE2C1AB. The two recombinants strains were subjected to fermentation and showed PHA accumulation, the first reported example of mcl-PHA production in B. subtilis. Gas Chromatography analysis identified the compound produced by B. subtilis/pBE2C1 to be a hydroxydecanoate-co-hydroxydodecanoate (HD-co-HDD) polymer whereas that produced by B. subtilis/pBE2C1AB was a hydroxybutyrate-co-hydroxyde-canoate-co-hydroxydodecanoate (HB-HD-HDD) polymer.     

Construction of the Pharmacophore Model of Glycogen Synthase Kinase-3 Inhibitors

A three dimensional pharmacophore model has been generated for glycogen synthase kinase-3 （GSK-3） inhibitots. A dataset consisting of 89 compounds was selected on the basis of the information content of the structures and activity data as required by the CATALYST system. The optimum model with four features （one hydrogen-bond acceptor unit, one ring aromatic unit, and two hydrophobic aromatic units） was selected with a good correlation coefficient （0.95）. This model is able to predict the activity of other known GSK-3 inhibitors not included in the model generation, and can be further used to identify structurally diverse compounds with desired biological activity by virtual screening.     

On The Fluorometric Method for the Measurement of β-Cyanoalanine Synthase Activity

Abstract

A modification of a fluorescence assay method for β-cyanoalanine synthase has been described which involves the direct coupling of the product of cyanide and cysteine to resazurin, to produce the fluorescent resosurfin, The fluorescence produced was found to be directly proportional to the rate of the reaction.     

A QM/MM study on the reaction pathway leading to 2‐Aceto‐2‐hydroxybutyrate in the catalytic cycle of AHAS

The reaction between the intermediate 2‐hydroxyethyl‐thiamin diphosphate (HEThDP^?) and 2‐ketobutyrate, in the third step of the catalytic cycle of acetodydroxy acid synthase, is addressed from a theoretical point of view by means of hybrid quantum/molecular mechanical calculations. The QM region includes one molecule of 2‐ketobutyrate, the HEThDP^? intermediate, and the residues Arg 380 y Glu 139; whereas the MM region includes the rest of the protein. The study includes potential energy surface scans to identify and characterize critical points on it, transition state search and activation barrier calculations. The results show that the reaction occurs via a two‐step mechanism corresponding to the carboligation and proton transfer in the first stage; and the product release in the second step. © 2014 Wiley Periodicals, Inc.