From a peptide lead to an orally active peptidomimetic fibrinogen receptor antagonist

Antagonists of the platelet fibrinogen receptor (GP IIb/IIIa receptor) are expected to be a new promising class of antithrombotic agents. The binding of fibrinogen to the fibrinogen receptor depends on an Arg-Gly-Asp-Ser (RGDS) tetrapeptide recognition motif. Structural modifications of the RGDS lead have led to the discovery of a non-peptide RGD mimetic GP IIb/IIIa antagonist 20 (S 1197). Compound 20 inhibits dose-dependently and reversibly human platelet aggregation. Modeling studies based on structure–activity data revealed the following structural features of the drug as important for receptor binding: the amidino group, the carboxylate group, hydrophobic substitutions at the carboxyl-terminus and at the side chain carrying the positive charge, the carboxyl-terminal NH group of the -amino acid as a hydrogen bond donor and one oxygen atom of the hydantoin as a hydrogen bond acceptor. The ethyl ester prodrug of 20 (S 5740) is an orally active antithrombotic agent which has the potential to be used to treat and prevent thrombotic diseases in humans.     

Restriction endonuclease EcoO109 from Escherichia coli H709c with heptanucleotide recognition site 5'-PuG/GNCCPy

A new restriction endonuclease, EcoO109, has been isolated from Escherichia coli H709c by polyethyleneimine (PEI) precipitation, DEAE-cellulose chromatography and heparin agarose chromatography. The yield was high, more than 3000 units/g of wet cells. The EcoO109 endonuclease recognizes and cleaves a nucleotide sequence of (formula: see text), in the presence of 10 mM Mg2+. The enzyme will be useful for structural analysis and molecular cloning of DNA because of the stability, high yield and easy handling of the producer strain.     

Ribosome specificity of archaebacterial elongation factor 2. Studies with hybrid polyphenylalanine synthesis systems

Polyphenylalanine synthesis with ribosomes and two separated, partially purified elongation factors (EF) was measured in cell-free systems from the archaebacteria Thermoplasma acidophilum and Methanococcus vannielii, in an eukaryotic system from rat liver and an eubacterial one with Escherichia coli ribosomes and factors from Thermus thermophilus. By substitution of heterologous EF-2 or EF-G, respectively, for the homologous factors, ribosome specificity was shown to be restricted to factors from the same kingdom. In contrast, EF-1 from T. thermophilus significantly cooperated with ribosomes from T. acidophilum.     

Different susceptibility of indigenous populations of Lymantria dispar to the exotic entomopathogen Entomophaga maimaiga

The recovery of the host‐specific entomopathogen Entomophaga maimaiga is still limited to certain world areas, although it is recently spreading to Eastern Europe. This study evaluated the effectiveness and fitness of an E. maimaiga isolate from Balkans against Lymantria dispar populations collected along the Italian peninsula and main islands, where the fungus has never been reported. As a result of different bioassays, the pathogenicity against gypsy moth larvae was generally confirmed, although significant differences among insects feeding upon diverse forest plant species were observed. The lack of significant susceptibility of other lepidopteran species from the same areas is also reported.     

Depletion of JMJD5 sensitizes tumor cells to microtubule-destabilizing agents by altering microtubule stability

Microtubules play essential roles in mitosis, cell migration, and intracellular trafficking. Drugs that target microtubules have demonstrated great clinical success in cancer treatment due to their capacity to impair microtubule dynamics in both mitotic and interphase stages. In a previous report, we demonstrated that JMJD5 associated with mitotic spindle and was required for proper mitosis. However, it remains elusive whether JMJD5 could regulate the stability of cytoskeletal microtubules and whether it affects the efficacy of microtubule-targeting agents. In this study, we find that JMJD5 localizes not only to the nucleus, a fraction of it also localizes to the cytoplasm. JMJD5 depletion decreases the acetylation and detyrosination of α-tubulin, both of which are markers of microtubule stability. In addition, microtubules in JMJD5-depleted cells are more sensitive to nocodazole-induced depolymerization, whereas JMJD5 overexpression increases α-tubulin detyrosination and enhances the resistance of microtubules to nocodazole. Mechanistic studies revealed that JMJD5 regulates MAP1B protein levels and that MAP1B overexpression rescued the microtubule destabilization induced by JMJD5 depletion. Furthermore, JMJD5 depletion significantly promoted apoptosis in cancer cells treated with the microtubule-targeting anti-cancer drugs vinblastine or colchicine. Together, these findings suggest that JMJD5 is required to regulate the stability of cytoskeletal microtubules and that JMJD5 depletion increases the susceptibility of cancer cells to microtubule-destabilizing agents.     

保肝降酶的天然药物研究进展

天然药物治疗肝损伤的功效及其低毒性已得到广泛的认可。由于大多肝损伤均有氧化应激的参与,天然药物的保肝作用通常与其抗氧化特性或激发体内抗氧化防御系统的能力有关。然而,越来越多的证据表明,除了抗氧化,天然药物还具有许多其他保肝机制。文章综述了近年来保肝降酶的天然药物的研究进展,将国内外已报道的天然药物分黄酮类、生物碱类、皂苷类、多糖类、木脂素类、萜类及其他类并分别列举其治疗机制及效果,为天然药物保肝降酶相关性的研究提供文献依据和研究思路,并对未来保肝降酶的新药研发方向进行展望。     

Twin anchors of the soybean isoflavonoid metabolon: evidence for tethering of the complex to the endoplasmic reticulum by IFS and C4H

Isoflavonoids are specialized plant metabolites, almost exclusive to legumes, and their biosynthesis forms a branch of the diverse phenylpropanoid pathway. Plant metabolism may be coordinated at many levels, including formation of protein complexes, or ‘metabolons’, which represent the molecular level of organization. Here, we have confirmed the existence of the long‐postulated isoflavonoid metabolon by identifying elements of the complex, their subcellular localizations and their interactions. Isoflavone synthase (IFS) and cinnamate 4–hydroxylase (C4H) have been shown to be tandem P450 enzymes that are anchored in the ER, interacting with soluble enzymes of the phenylpropanoid and isoflavonoid pathways (chalcone synthase, chalcone reductase and chalcone isomerase). The soluble enzymes of these pathways, whether localized to the cytoplasm or nucleus, are tethered to the ER through interaction with these P450s. The complex is also held together by interactions between the soluble elements. We provide evidence for IFS interaction with upstream and non‐consecutive enzymes. The existence of such a protein complex suggests a possible mechanism for flux of metabolites into the isoflavonoid pathway. Further, through interaction studies, we identified several candidates that are associated with GmIFS2, an isoform of IFS, in soybean hairy roots. This list provides additional candidates for various biosynthetic and structural elements that are involved in isoflavonoid production. Our interaction studies provide valuable information about isoform specificity among isoflavonoid enzymes, which may guide future engineering of the pathway in legumes or help overcome bottlenecks in heterologous expression.     

Poly (l-lysine) based semi-interpenetrating polymer network as pH-responsive hydrogel for controlled release of a model protein drug streptokinase

With the aim of developing a pH-sensitive controlled drug release system, a poly (L-lysine) (PLL) based cationic semi-interpenetrating polymer network (semi-IPN) has been synthesized. This cationic hydrogel was designed to swell at lower pH and de-swell at higher pH and therefore be applicable for achieving regulated drug release at a specific pH range. In addition to the pH sensitivity, this hydrogel was anticipated to interact with an ionic drug, providing another means to regulate the release rate of ionic drugs. This semi-IPN hydrogel was prepared using a free-radical polymerization method and by crosslinking of the polyethylene glycol (PEG)-methacrylate polymer through the PLL network. The two polymers were penetrated with each other via interpolymer complexation to yield the semi-IPN structures. The PLL hydrogel thus prepared showed dynamic swelling/de-swelling behavior in response to pH change, and such a behavior was influenced by both the concentrations of PLL and PEG-methacrylate. Drug release from this semi-IPN hydrogel was also investigated using a model protein drug, streptokinase. Streptokinase release was found to be dependent on its ionic interaction with the PLL backbones as well as on the swelling of the semi-IPN hydrogel. These results suggest that a PLL semi-IPN hydrogel could potentially be used as a drug delivery platform to modulate drug release by pH-sensitivity and ionic interaction.     

Structural basis of substrate specificity in the serine proteases.

Structure-based mutational analysis of serine protease specificity has produced a large database of information useful in addressing biological function and in establishing a basis for targeted design efforts. Critical issues examined include the function of water molecules in providing strength and specificity of binding, the extent to which binding subsites are interdependent, and the roles of polypeptide chain flexibility and distal structural elements in contributing to specificity profiles. The studies also provide a foundation for exploring why specificity modification can be either straightforward or complex, depending on the particular system.     

Limitation of net CO2 assimilation rate by internal resistances to CO2 transfer in the leaves of two tree species (Fagus sylvatica L. and Castanea sativa Mill.)

Using a combination of gas-exchange and chlorophyll fluorescence measurements, low apparent CO₂/O₂ specificity factors (1300 mol mol_?1) were estimated for the leaves of two deciduous tree species (Fagus sylvatica and Castanea sativa). These low values contrasted with those estimated for two herbaceous species and were ascribed to a drop in the CO₂ mole fraction between the intercellular airspace (C_i) and the catalytic site of Rubisco (C_c) due to internal resistances to CO₂ transfer. C_c. was calculated assuming a specificity of Rubisco value of 2560 mol mol^?1. The drop between C_i and C_c was used to calculate the internal conductance for CO₂ (g_i). A good correlation between mean values of net CO₂ assimilation rate (A) and g_i was observed within a set of data obtained using 13 woody plant species, including our own data. We report that the relative limitation of A, which can be ascribed to internal resistances to CO₂ transfer, was 24–30%. High internal resistances to CO₂ transfer may explain the low apparent maximal rates of carboxylation and electron transport of some woody plant species calculated from A/C_i curves.