Influence of increasing D/A ratio on geometric and electronic properties of D–A-type copolymers

A series of D–A-type copolymers was designed and studied systematically for the purpose of gaining a deeper understanding of how the D/A ratio may influence the geometric and electronic properties when it varies from 2:1 to 6:1 by using DFT method. The results show that it has a significant effect on the bond length alternation, band gap, bandwidth and effective mass of carriers of the designed D–A-type copolymers. But its influence on the geometric and electronic properties shows something very different in degree when it increases from 2:1 to 4:1 and then increases further from 4:1 to 6:1. It is found that the effects of increasing D/A ratio on geometric and electronic properties are much stronger when the D/A ratio increases from 2:1 to 4:1 than when it further increases from 4:1 to 6:1. The theoretical results show that the polymers with a D/A ratio of 2:1 have much smaller effective mass of carriers and much wider bandwidth compared to those polymers with the D/A ratio of 4:1 and 6:1. Therefore, the designed D–A-type copolymers with a D/A ratio of 2:1 may actually be the better candidates for intrinsic conduction materials.     

Elite swimmers and the D allele of the ACE I/D polymorphism

A polymorphism of the human angiotensin-1-converting enzyme (ACE) gene has been identified in which the presence (insertion, I allele) of a 287-bp fragment rather than the absence (deletion, D allele) is associated with lower ACE activity. Several recent studies have shown an association of the I allele with endurance performance, it being found with excess frequency in elite distance runners, rowers and mountaineers. Other workers using heterogeneous cohorts of athletes from mixed sporting disciplines have found no such association. An increasing linear trend of I allele frequency with the distance run amongst Olympic runners and an excess of the D allele amongst sprinters led us to examine whether the ratio of I and D alleles in swimmers competing over different distances would also vary. Swimmers (n=120) from the European and Commonwealth championships and an American college team had their ACE genotype determined and their gene and allele frequencies compared with several control groups, the most closely age-matched of which were 1,248 military recruits. Of the 103 Caucasians, there was a significant excess of the D allele compared with this control group only in the truly elite swimmers of the European and Commonwealth championships (P=0.004). This association remained in those competing over shorter distances (P=0.005 for 400 m and below) but not in the longer events. These findings were confirmed in three further large control groups. A population association study testing whether a genetic marker (the ACE I/D polymorphism) occurs more frequently in cases (elite athletes) than in controls therefore requires a homogeneous cohort of subjects from the same sporting discipline.     

CD Spectra of D1/D2/Cyt b559 Complax in Photodamaged Photosystem Ⅱ Reaction Center

The CD spectrum of photosystem Ⅱ reaction center D1/D2/Cyt b559 complex showed a strong reverse band with positive peak at 680 nm and negative peak at 660 nm in the red absorption region (Qy band). After the D1/D2/Cyt b559 complex was illuminated by strong light, the CD signals of the complex decreased significantly in the red region in which the negative peak still existed but the positive one disappeared. The result suggested that the CD signal of photosystem Ⅱ reaction center D1/D2/Cyt b559 complex not only came from the primary donor, P680, but also from other pigments such as from accessory Chl a or Pheo a.     

Effects of Chronic Administration of Interferon α A/D on Serotonergic Receptors in Rat Brain

The effects of chronic administration of interferon (IFN; recombinant human IFN -A/D) on serotonergic binding sites in rat brain were investigated. IFN was injected daily for 2 weeks at a dose of 100000 I.U./kg, (i.p.) in male Wistar rats. IFN did not alter either [³H]ketanserin binding to 5-HT_2A receptors or [³H]paroxetine binding to 5-HT transporters. Scatchard analysis of [³H]8-hydroxy-dipropylaminotetraline (8-OH-DPAT) binding to 5-HT_1A receptors demonstrated the presence of high- and low-affinity binding sites in both treatment and control groups. IFN significantly increased both Kd and Bmax measures of [³H]8-OH-DPAT binding at low-affinity binding sites, but not at the high-affinity sites. These results suggest that IFN affects the low-affinity 5-HT_1A receptors sites and may be involved in the development of IFN-induced psychiatric disturbances.     

Involvement of RhoA/ROCK in insulin secretion of pancreatic β-cells in 3D culture

Cell-cell contacts and interactions between pancreatic β-cells and/or other cell populations within islets are essential for cell survival, insulin secretion, and functional synchronization. Three-dimensional (3D) culture systems supply the ideal microenvironment for islet-like cluster formation and functional maintenance. However, the underlying mechanisms remain unclear. In this study, mouse insulinoma 6 (MIN6) cells were cultured in a rotating 3D culture system to form islet-like aggregates. Glucose-stimulated insulin secretion (GSIS) and the RhoA/ROCK pathway were investigated. In the 3D-cultured MIN6 cells, more endocrine-specific genes were up-regulated, and GSIS was increased to a greater extent than in cells grown in monolayers. RhoA/ROCK inactivation led to F-actin remodeling in the MIN6 cell aggregates and greater insulin exocytosis. The gap junction protein, connexin 36 (Cx36), was up-regulated in MIN6 cell aggregates and RhoA/ROCK-inactivated monolayer cells. GSIS dramatically decreased when Cx36 was knocked down by short interfering RNA and could not be reversed by RhoA/ROCK inactivation. Thus, the RhoA/ROCK signaling pathway is involved in insulin release through the up-regulation of Cx36 expression in 3D-cultured MIN6 cells.     

Light-Induced Damage of Pheophytin a Molecule in the Isolated Photosystem Ⅱ Reaction Center D1/D2/Cyt b559 Complex

Photodamage of some pigments in the isolated photosystem Ⅱ (PS Ⅱ ) reaction center D1/D2/Cyt b559 complex from spinach has been investigated by means of high performance liquid chromatography. The light-induced damage of pheophytin a (pheo a) in the complex was observed for the first time. The content of pheo a decreased about 47 % by illumination, suggesting only one of the two pheo a molecules in the PS Ⅱ reaction center complex was damaged. No damage of β-carotene was found.     

D1/D2/Cytb-559复合物的共振拉曼光谱

光系统Ⅱ（ＰＳⅡ）反应中心Ｄ１／Ｄ２／Ｃｙｔｂ－５５９ 复合物在４８８ ｎｍ 处激发的共振拉曼光谱显示４ 个主要谱带,其峰位分别在１５３２（υ１）、１１６５（υ２）、１０１０（υ３）和９７０（υ４） ｃｍ － １处,表明ＰＳⅡ反应中心结合的β－胡萝卜素分子是全反式构型。Ｄ１／Ｄ２／Ｃｙｔｂ－５５９ 复合物的色素抽提液的拉曼光谱也显示４ 个主要的拉曼峰,其中υ４ 谱带的强度急剧下降,说明ＰＳⅡ反应中心内部结合的β－胡萝卜素分子与抽提液中自由的β－胡萝卜素分子的构象不同,而与光合细菌反应中心内部的类胡萝卜素分子的构象相似,其共轭多烯链的平面也处于扭曲状态     

Structure-guided development of dual β2 adrenergic/dopamine D2 receptor agonists

Aiming to discover dual-acting β₂ adrenergic/dopamine D₂ receptor ligands, a structure-guided approach for the evolution of GPCR agonists that address multiple targets was elaborated. Starting from GPCR crystal structures, we describe the design, synthesis and biological investigation of a defined set of compounds leading to the identification of the benzoxazinone (R)-3, which shows agonist properties at the adrenergic β₂ receptor and substantial G protein-promoted activation at the D₂ receptor. This directed approach yielded molecular probes with tuned dual activity. The congener desOH-3 devoid of the benzylic hydroxyl function was shown to be a β₂ adrenergic antagonist/D₂ receptor agonist with K_i values in the low nanomolar range. The compounds may serve as a promising starting point for the investigation and treatment of neurological disorders.     

The Resonance Raman Spectra of β-Carotene Molecules in the Photosystem Ⅱ Reaction Center D1/D2/Cyt b-559 Complex

The resonance Raman spectrum of β-carotene in photosystem Ⅱ (PS Ⅱ )reaction center complex was characterized by four main bands, peaking at 1532 (νl), 1156 (ν2), 1010 (ν3) and 970 (ν4) cm -1, respectively, with several additional small Raman bands in the region between 1100 cm-1 and 1500 cm-1 It was suggested that β-carotene molecules of the reaction center complex were in all-trans configuration. The resonance Raman spectrum of an acetone extract from the reaction center complex also showed four main bands. The peak position of νl, ν3 and ν4 band shifted 5 cm-1 to the shorter wave number. The most dramatic changes were the reduction of the intensity of ν4. From the above results it was demon- strated that the conformation of β-carotene molecules in the PS Ⅰ reaction center was not the same as that of free β-carotene molecules in solution, but similar to that of carotenoid molecules in the photosynthetic bacterial reaction center, in other words, they are likely to be in a twisted conformation.     

β-catenin/cyclin D1 mediated development of suture mesenchyme in calvarial morphogenesis

Background  

Mouse genetic study has demonstrated that Axin2 is essential for calvarial development and disease. Haploid deficiency of β-catenin alleviates the calvarial phenotype caused by Axin2 deficiency. This loss-of-function study provides evidence for the requirement of β-catenin in exerting the downstream effects of Axin2.     

Staphylococcus aureusα-Toxin: Characterization of Protein/Lipid Interactions, 2D Crystallization on Lipid Monolayers, and 3D Structure

Staphylococcus aureusα-toxin was characterized with respect to surface activity and its interaction with lipid monolayers. The protein alone had a detergent-like behavior at the air/water interface. Its affinity was higher for negatively charged than for neutral phospholipids. The interaction was pH dependent, showing a maximum increase at pH 7.0. Only a small part of the protein oligomer appeared to be inserted into the monolayers. Crystalline sheets of α-toxin were formed using negatively charged phospholipids. Electron microscopy of such areas, at different tilt angles, allowed reconstruction of a three-dimensional model following image processing. The sheets analyzed consisted of two protein layers arranged on a tetragonal lattice. Under the conditions used to grow the crystals the toxin formed 90-Å-wide cylinders with a height of 70 Å. One of the imposed fourfold axes running perpendicular to the plane of the crystalline layer is positioned at a protein-deficient region which forms a 25-Å-wide pore through the oligomer.     

Function of two β-carotenes near the D1 and D2 proteins in photosystem II dimers

The antenna proteins in photosystem II (PSII) not only promote energy transfer to the photosynthetic reaction center (RC) but provide also an efficient cation sink to re-reduce chlorophyll a if the electron transfer (ET) from the Mn-cluster is inhibited. Using the newest PSII dimer crystal structure (3.0 Å resolution), in which 11 β-carotene molecules (Car) and 14 lipids are visible in the PSII monomer, we calculated the redox potentials (E_m) of one-electron oxidation for all Car (E_m(Car)) by solving the Poisson-Boltzmann equation. In each PSII monomer, the D1 protein harbors a previously unlocated Car (Car_D1) in van der Waals contact with the chlorin ring of Chl_Z(D1). Each Car_D1 in the PSII dimer complex is located in the interface between the D1 and CP47 subunits, together with another four Car of the other PSII monomer and several lipid molecules. The proximity of Car bridging between Car_D1 and plastoquinone/Q_A may imply a direct charge recombination of Car⁺Q_A⁻. The calculated E_m(Car_D1) and E_m(Chl_Z(D1)) are, respectively, 83 and 126 mV higher than E_m(Car_D2) and E_m(Chl_Z(D2)), which could explain why Car_D2⁺ and Chl_Z(D2)⁺ are observed rather than the corresponding Car_D1⁺ and Chl_Z(D1)⁺.     

Orientation of Pigments in the Isolated Photosystem ￠ò Sub-core Reaction Center CP47/D1/D2/Cyt b-559 Complexes: A Linear Dichrosism Study

Linear dichroism (LD) spectroscopy is an important technique in the study of the orientation and organization of pigments in the photosynthetic membrane complexes in vivo and in vitro. In this work, the orientation of the pigments in the isolated photosystem Ⅱ (PSⅡ) sub-core reaction center complexes was analyzed and characterized by means of low temperature absorption and LD spectroscopy. The preparations containing different amounts of CP47 isolated from spinach (Spinacia oleracea L.) chloroplast were used in order to investigate the orientation of pigments in the PSⅡ sub-core CP47/D1/D2/Cyt b-559 (CP47/D1/D2) complexes. Chlorophyll a (Chl a) absorbing at 680 nm in CP47/D1/D2/Cyt b-559 complex showed an orientation of the Q y transition parallel to the membrane plane. It is proposed that there are two forms of β-carotene (β-Car) in CP47/D1/D2/Cyt b-559 complex, denoted as β-Car (Ⅰ)and β-Car (Ⅱ), with different orientations, β-Car (Ⅰ) at 470 and 505 nm is roughly parallel to the membrane plane, and β-Car (Ⅱ) at 460 and 490 nm seems to be perpendicular orientation. Upon the photoinhibitory experiment β-Car (Ⅱ) was found to be photosensitive and easily photodamaged. It also showed that the positive LD signal observed at 680 nm was quite complicated. This signal is tentatively attributed to P680 and some Chl　a of antenna in CP47 protein based upon our measurements.     

Validation of the BacT/ALERT?3D automated culture system for the detection of microbial contamination of epithelial cell culture medium

Living tissue engineering for regenerative therapy cannot withstand the usual pharmacopoeia methods of purification and terminal sterilization. Consequently, these products must be manufactured under aseptic conditions at microbiologically controlled environment facilities. This study was proposed to validate BacT/ALERT(?)3D automated culture system for microbiological control of epithelial cell culture medium (ECCM). Suspensions of the nine microorganisms recommended by the European Pharmacopoeia (Chap. 2.6.27: "Microbiological control of cellular products"), plus one species from oral mucosa and two negative controls with no microorganisms were prepared in ECCM. They were inoculated in FA (anaerobic) and SN (aerobic) culture bottles (Biomérieux, Lyon, France) and incubated in a BacT/ALERT(?)3D automated culture system. For each species, five sets of bottles were inoculated for reproducibility testing: one sample was incubated at the French Health Products Agency laboratory (reference) and the four others at Cell and Tissue Bank of Lyon, France. The specificity of the positive culture bottles was verified by Gram staining and then subcultured to identify the microorganism grown. The BacT/ALERT(?)3D system detected all the inoculated microorganisms in less than 2 days except Propionibacterium acnes which was detected in 3 days. In conclusion, this study demonstrates that the BacT/ALERT(?)3D system can detect both aerobic and anaerobic bacterial and fungal contamination of an epithelial cell culture medium consistent with the European Pharmacopoeia chapter 2.6.27 recommendations. It showed the specificity, sensitivity, and precision of the BacT/ALERT(?)3D method, since all the microorganisms seeded were detected in both sites and the uncontaminated medium ECCM remained negative at 7 days.     

The synthesis of 6,9-deepoxy-6,9-N-phenylimino-Δ6,8-prostaglandin I1 a novel inhibitor of leukotriene C and D synthesis

The pyrrole analog of prostacyclin, 6,9-deepoxy-6,9-N-phenylimino-Δ^6,8-prostaglandin I₁ was synthetized from PGF₂α methyl ester. This pyrroloprostacyclin (U-60, 257) and its methyl ester (U-56, 467) have been shown to inhibit leukotriene C/D biosynthesis and antagonized leukotriene C/D contractions in vitro. Antigen induced bronchopulmonary changes in monkeys and guinea pigs are inhibited by U-60, 257 in vivo.     

Light-induced Damage of Photosystem Ⅱ Primary Electron Donor P680: a High Performance Liquid Chromatographic Analysis of Pigment Content in D1/D2/Cytochrome b559 Complex Under Photoinhibitory Conditions

By HPLC analytical method, the change of PS Ⅱ RC' s pigment content in the process of photodamage under strong illumination from spinach ( Spinacia oleracea Mill. ) was comparatively studied. The experimental results show that: (1) In authors' analytical conditions, (of which, [Chl] = 150 µg/mL, and the illumination strength was put at 2.3 ×10 6 mJ·m－2·s－1 ), 45 rain of illumination could cause almost the whole loss of A680 in the fourth derivative absorption spectra, while A670 decreased to about one half of its original intensity; the absorption maximum in red, concurrently, was shifted from 676 nm to 671 nm, representing the loss of more than 90% of the photochemical activities of the PS Il RC. (2) During the period of continuous illumination, the Chl concentration decreased in a 3-period style, which meant that the first [Chl] decreased to the 2/3 of its original amount from 20 min to 40 rain after illumination had started, then became stabilized up to about 60 min of illumination, there after a second decrease of [ Chl ] in another about 20 min until it reached about 30 % of the original level and remained unchanged from about 80 min on. The original pigment components of D1/D2/Cyt b559 was approximately as 6 Chl a:2 Pheo:2β-Car which are in support of authors' previous proposal about the minimum Chl/Pheo ratio of 4: 2 in PS Ⅱ RC’s pigment contents. (3) After about 40 min of illumination, a newly appeared elution peak was found between the Pheo andβ-Car peaks in HPLC profile, at the retention time of 7.2 min, a little later than that (6.9 rain) of Pheo molecules, the newly appeared elution peak was supposed to be a kind of accumulated and stable product of the PS II RC's photodamage process and very much possible the Pheo-like molecules.     

Palmitoylation Regulates Intracellular Trafficking of β(2) Adrenergic Receptor/Arrestin/Phosphodiesterase 4D Complexes in Cardiomyocytes

β(2) adrenergic receptor (β(2)AR) is a prototypical G-protein coupled receptor that stimulates the classic cAMP-protein kinase A (PKA) signaling pathway. Recent studies indicate that the cAMP-PKA activities are spatiotemporally regulated in part due to dynamic association of β(2)AR with phosphodiesterase 4D (PDE4D), a group of cAMP degradation enzymes. Here, we demonstrate that in cardiomyocytes, palmitoylation of β(2)AR, the covalent acylation of cysteine residue 341, plays a critical role in shaping subcellular cAMP-PKA activities in cardiomyocytes via regulating β(2)AR association with arrestin/PDE4D. Replacing cysteine 341 on β(2)AR with alanine (C341A) leads to an impaired binding to β arrestin 2. Surprisingly, the C341A mutant is able to internalize via an arrestin-independent pathway at saturated concentration of agonist stimulation; the internalization becomes caveolae-dependent and requires dynamin GTPase. However, the impaired binding to β arrestin 2 also leads to an impaired recruitment of PDE4D to the C341A mutant. Thus, the mutant C341A β(2)AR is transported alone from the plasma membrane to the endosome without recruiting PDE4D. This alteration leads to an enhanced cytoplasmic cAMP signal for PKA activation under β(2)AR stimulation. Functionally, Mutation of the C341 residue or inhibition of palmitoylation modification of β(2)AR enhances the receptor-induced PKA activities in the cytoplasm and increases in myocyte contraction rate. Our data reveal a novel function of palmitoylation in shaping subcellular cAMP-PKA signaling in cardiomyocytes via modulating the recruitment of β arrestin 2-PDE4D complexes to the agonist-stimulated β(2)AR.     

Observation of One Pheophytin a Molecule Not Associated with Primary Photochemistry in the Isolated Photosystem Ⅱ Reaction Center D1/D2/Cyt b559 Complex

Photodamage of pheophytin a (pheo a) in the isolated photosystem Ⅱ (PSⅡ ) reaction center D1/D2/Cyt b559 complex from spinach has been investigated by high performance liquid chromatographic method in detail. The results showed that: (1) There is one pheo a molecule which is not associated with the primary photochemistry in the PS Ⅱ reaction center complex. It may be considered that there are two different electron transfer branches in the PS Ⅱ reaction center just as in the purple bacterium photosynthetic reaction center. (2) The damaged pheo a may be attributed to the one bonding to the D2 protein comparing the D2 subunit in the PS Ⅱ reaction center with M subunit in the purple bacterium photosynthetic reaction center. (3) A possible arrangement model of redox cofactors in the PS Ⅱ reaction center was proposed based on our experiment.     

Cloning and characterization of the bgxA gene from Erwinia chrysanthemi D1 which encodes a β-glucosidase/xylosidase enzyme

A -glucosidase/xylosidase gene from Erwinia chrysanthemi strain D1 was cloned and sequenced. This gene, named bgxA, encodes a ca. 71 kDa protein product which, following removal of the leader peptide, resulted in a ca. 69 kDa mature protein that accumulated in the periplasmic space of E. chrysanthemi strain D1 and Escherichia coli cells expressing the cloned gene. The protein exhibited both -glucosidase and -xylosidase activities but gave no detectable activity on xylan or carboxymethyl cellulose. The enzyme was classified as a type 3 glycosyl hydrolase, but was unusual in having a truncated B region at the carboxyl-terminus. Several E. chrysanthemi strains isolated from corn produced the glucosidase/xylosidase activity but not those isolated from dicot plants. However, bgxA marker exchange mutants of strain D1 were not detectably altered in virulence on corn leaves.