Comparison of Anti-Hyperglycemic Effect of Inorganic Constituents and Organic in Traditional Chinese Medicine,Jinqi Compound Recipe

An attempt has been made to compare the hypoglycemic effect of the organic constituents and the inorganic constituents in traditional Chinese medicine, Jinqi compound recipe. Alloxan-induced hyperglycemic mice were used in the study. The body weights, blood glucose, HbA1c, insulin secretion, and glycogen synthesis of the mice were analyzed respectively. After the mice were administered (oral) with organic constituents, the blood glucose and the HbA1c of alloxan-induced hyperglycemic mice decreased (p < 0.05, p < 0.01) and the glycogen synthesis of alloxan-induced hyperglycemic mice elevated (p < 0.05). Also, the body weight of the alloxan-induced hyperglycemic mice was increased gradually. However, the same result did not occur in the inorganic constituent-treated groups. It is noted that neither organic constituents nor inorganic constituents could elevate the level of serum insulin in the alloxan-induced hyperglycemic mice significantly. It is implied that the hypoglycemic effect for type 2 diabetes was caused by the organic constituents of Jinqi recipe. The results can be used to set new standards to control the quality of the Jinqi recipe.     

The content of total sulphur and sulphur forms in birch (Betula pendula Roth) leaves in the air-polluted Krusne hory mountains

In leaves of birch (Betula pendula Roth), changes in the content of total sulphur and its inorganic and organic forms were determined in relation to the decreasing air-pollution load (SO₂) in the air-polluted Krusne hory mountains and the Decin sandstone highlands in 1995, 1998, 2001 and 2004. Results have shown that birch is able to use considerable amounts of sulphur taken through leaves from air-pollution load. Birch responds fast to changes in air-pollution load by fall in the content of total and inorganic forms of sulphur in leaves.     

Development of a charcoal paper adenosine triphosphate:pyrophosphate exchange assay: Kinetic characterization of NEDD8 activating enzyme

Ubiquitin activating enzyme (UAE, UBE1, or E1) and seven known homologous “E1s” initiate the conjugation pathways for ubiquitin and 16 other ubiquitin-like modifiers (ULMs) found in humans. The initial step catalyzed by E1s uses adenosine triphosphate (ATP) to adenylate the C terminus of the appropriate ULM and results in the production of inorganic pyrophosphate (PPi). The mechanism of these enzymes can be studied with assays that measure the rate of ULM-dependent ATP:PPi exchange. The traditional method follows the initial velocity of [³²P]PPi incorporation into ATP by capturing the nucleotide on activated charcoal powder to separate it from excess [³²P]PPi and then measuring [³²P]ATP in a scintillation counter. We have modified the method by using charcoal paper to capture the nucleotide and a phosphorimager to quantify the [³²P]ATP. The significant increase in throughput that these modifications provide is accomplished without any sacrifice in sensitivity or accuracy compared with the traditional method. To demonstrate this, we reproduce and extend the characterization of the NEDD8 activating enzyme.     

Role of polyphosphate in regulation of the Streptomyces lividans KcsA channel

We examine the hypotheses that the Streptomyces lividans potassium channel KcsA is gated at neutral pH by the electrochemical potential, and that its selectivity and conductance are governed at the cytoplasmic face by interactions between the KcsA polypeptides and a core molecule of inorganic polyphosphate (polyP). The four polypeptides of KcsA are postulated to surround the end unit of the polyP molecule with a collar of eight arginines, thereby modulating the negative charge of the polyP end unit and increasing its preference for binding monovalent cations. Here we show that KcsA channels can be activated in planar lipid bilayers at pH 7.4 by the chemical potential alone. Moreover, one or both of the C-terminal arginines are replaced with residues of progressively lower basicity-lysine, histidine, valine, asparagine-and the effects of these mutations on conductance and selectivity for K⁺ over Mg²⁺ is tested in planar bilayers as a function of Mg²⁺ concentration and pH. As the basicity of the C-terminal residues decreases, Mg²⁺ block increases, and Mg²⁺ becomes permeant when medium pH is greater than the pI of the C-terminal residues. The results uphold the premise that polyP and the C-terminal arginines are decisive elements in KcsA channel regulation.     

福氏痢疾杆菌组氨酸合成中双功能焦磷酸水解酶和磷酸核糖环化水解酶双功能蛋白的初步研究

福志贺氏菌属是一类革兰氏阴性杆菌,是引起人类细菌性痢疾的主要致病源。以福志贺氏痢疾杆菌2a型301株全基因组为模板、以pET22b-(+)为载体、克隆了一个关键基因：组氨酸合成途径中双功能焦磷酸水解酶和磷酸核糖环化水解酶蛋白（简称：sf2088）。以BL21（DE3）为表达菌株,优化表达条件,获得了可溶表达的蛋白。经过亲和层析和分子筛层析获得目标蛋白。采用分析超速离心和动态光散射实验,对纯化后的蛋白进行稳定聚集状态的条件搜索,结果发现锌离子对稳定其聚合状态很重要。通过正交实验,获得蛋白保持稳定聚集态的条件。这对该酶的进一步研究奠定了基础。     

Crystal structures of human NUDT5 reveal insights into the structural basis of the substrate specificity

Human NUDT5 (hNUDT5) is an ADP-ribose pyrophosphatase (ADPRase) belonging to the Nudix hydrolase superfamily. It presumably plays important roles in controlling the intracellular level of ADP-ribose (ADPR) to prevent non-enzymatic ADP-ribosylation by hydrolyzing ADPR to AMP and ribose 5'-phosphate. We report here the crystal structures of hNUDT5 in apo form, in complex with ADPR, and in complex with AMP with bound Mg2+. hNUDT5 forms a homodimer with substantial domain swapping and assumes a structure more similar to Escherichia coli ADPRase ORF209 than human ADPRase NUDT9. The adenine moiety of the substrates is specifically recognized by the enzyme via hydrogen-bonding interactions between N1 and N6 of the base and Glu47 of one subunit, and between N7 of the base and Arg51 of the other subunit, providing the molecular basis for the high selectivity of hNUDT5 for ADP-sugars over other sugar nucleotides. Structural comparisons with E. coli ADPRase ORF209 and ADPXase ORF186 indicate that the existence of an aromatic residue on loop L8 in ORF186 seems to be positively correlated with its enzymatic activity on APnA, whereas hNUDT5 and ORF209 contain no such residue and thus have low or no activities on APnA.     

Colorimetric inorganic pyrophosphate assay using a double cycling enzymatic method

Pyruvate phosphate dikinase (PPDK, EC 2.7.9.1) from the hyperthermophile Thermotoga maritima was biochemically characterized with the aim of establishing a colorimetric assay for inorganic pyrophosphate (PPi). When heterologously expressed in Escherichia coli, T. maritima PPDK (TmPPDK) was far more stable any other PPDK reported so far: it retained >90% of its activity after incubation for 1 h at 80 °C, and >80% of its activity after incubation for 20 min at pHs ranging from 6.5 to 10.5 (50 °C). In contrast to PPDKs from protozoa and plants, this TmPPDK showed very long-term stability at low temperature: full activity was retained even after storage for at least 2 years at 4 °C. TmPPDK was successfully applied to a novel colorimetric PPi assay, which employed (i) a PPi cycling reaction using TmPPDK and nicotinamide mononucleotide adenylyltransferase (EC 2.7.7.1) from Saccharomyces cerevisiae and (ii) a NAD cycling reaction to accumulate reduced nitroblue tetrazolium (diformazan). This enabled detection of 0.2 μM PPi, making this method applicable for preliminary measurement of PPi levels in PCR products in an automatic clinical analyzer.     

Exploring emergent properties in cellular homeostasis using OnGuard to model K and other ion transport in guard cells

It is widely recognized that the nature and characteristics of transport across eukaryotic membranes are so complex as to defy intuitive understanding. In these circumstances, quantitative mathematical modeling is an essential tool, both to integrate detailed knowledge of individual transporters and to extract the properties emergent from their interactions. As the first, fully integrated and quantitative modeling environment for the study of ion transport dynamics in a plant cell, OnGuard offers a unique tool for exploring homeostatic properties emerging from the interactions of ion transport, both at the plasma membrane and tonoplast in the guard cell. OnGuard has already yielded detail sufficient to guide phenotypic and mutational studies, and it represents a key step toward ‘reverse engineering’ of stomatal guard cell physiology, based on rational design and testing in simulation, to improve water use efficiency and carbon assimilation. Its construction from the HoTSig libraries enables translation of the software to other cell types, including growing root hairs and pollen. The problems inherent to transport are nonetheless challenging, and are compounded for those unfamiliar with conceptual ‘mindset’ of the modeler. Here we set out guidelines for the use of OnGuard and outline a standardized approach that will enable users to advance quickly to its application both in the classroom and laboratory. We also highlight the uncanny and emergent property of OnGuard models to reproduce the ‘communication’ evident between the plasma membrane and tonoplast of the guard cell.