Nicotinamidase is involved in the maintenance of NAD+ homeostasis and in the NAD+ salvage pathway of most prokaryotes, and it is considered as a possible drug target. The gene (ASAC_0847) encoding a hypothetical nicotinamidase has been found in the genome of the thermophilic archaeon Acidilobus saccharovorans. The product of this gene, NA_As0847, has been expressed in Escherichia coli, isolated, and characterized as a Fe2+-containing nicotinamidase (kcat/Km = 427 mM?1·sec?1)/pyrazinamidase (kcat/Km = 331 mM?1·sec?1). NA_As0847 is a homodimer with molecular mass 46.4 kDa. The enzyme has high thermostability (T1/2 (60°C) = 180 min, T1/2 (80°C) = 35 min) and thermophilicity (Topt = 90°C, Ea = 30.2 ± 1.0 kJ/mol) and broad pH interval of activity, with the optimum at pH 7.5. Special features of NA_As084 are the presence of Fe2+ instead of Zn2+ in the active site of the enzyme and inhibition of the enzyme activity by Zn2+ at micromolar concentrations. Analysis of the amino acid sequence revealed a new motif of the metal-binding site (DXHXXXDXXEXXXWXXH) for homological archaeal nicotinamidases. 相似文献
Lobophorins A (1) and B (2) belong to a large group of spirotetronate natural products with potent antibacterial and antitumor activities. The cloning of the lobophorin biosynthesis gene cluster from the deep-sea-derived Streptomyces sp. SCSIO 01127 identified a sugar-O-methyltransferase-encoding gene lobS1. The lobS1 inactivation mutant accumulated two new lobophorin analogs 3 and 4, different from 1 and 2 by lacking the 4-methyl group at the terminal l-digitoxose, respectively. Biochemical experiments verified that LobS1 was a SAM-dependent sugar-O-methyltransferase that required divalent metal ions for better activity. Antibacterial assays revealed compounds 3 and 4 were generally less potent than compounds 1 and 2. These findings suggest that the methylation on the terminal digitoxose by LobS1 tailors lobophorin biosynthesis and highlights the importance of this methylation for antibacterial potence. 相似文献
The molecular mechanism of potassium ion transport across membranes in conifers is poorly known. We isolated and analyzed a gene encoding a potassium transporter from the coniferCryptomeriajaponica.
Abstract
Potassium ion (K+) is an essential and the most abundant intracellular cation in plants. The roles of K+ in various aspects of plant life are closely linked to its transport across biological membranes such as the plasma membrane and the tonoplast, which is mediated by membrane-bound transport proteins known as transporters and channels. Information on the molecular basis of K+ membrane transport in trees, especially in conifers, is currently limited. In this study, we isolated one complementary DNA, CjKUP1, which is homologous to known plant K+ transporters, from Cryptomeriajaponica. Complementation tests using an Escherichiacoli mutant, which is deficient in K+ uptake activity, was conducted to examine the K+ uptake function of the protein encoded by CjKUP1. Transformation of the K+-uptake-deficient mutant with CjKUP1 complemented the deficiency of this mutant. This result indicates that CjKUP1 has a function of K+ uptake. The expression levels of CjKUP1 in male strobili were markedly higher from late September to early October than in other periods. The expression levels in male and female strobili were higher than those in other organs such as needles, inner bark, differentiating xylem, and roots. These results indicate that CjKUP1 is mainly involved in K+ membrane transport in the cells of reproductive organs of C. japonica trees, especially in male strobili during pollen differentiation. 相似文献
Altered numbers and functions of T cells have previously been demonstrated in chronic lymphocytic leukemia (CLL) patients. However, dynamics and specific T-cell subset alterations have not been studied in great detail. Therefore, we studied CLL blood lymphocyte subsets of individual patients in a longitudinal manner. Dynamic expansions of blood CD4+ and CD8+ T-cell numbers were consistently associated with a progressively increasing CLL leukemic compartment. Interestingly, the T-cell subset expansion over time was more pronounced in CD38+ CLL. Additionally, we performed gene expression profiling of CD3+ T cells of CLL patients and normal donors. Using gene set enrichment analysis, we found significant enrichment of genes with higher expression in CLL T cells within CD8+ effector memory and terminal effector T-cell gene signatures. In agreement with these data, we observed a marked expansion of phenotypic CD8+ effector memory T cells in CLL by flow cytometry. Moreover, we observed that increments of CD8+ effector memory T cells in human CLL and also mouse CLL (Eμ-TCL1 model) were due to an expansion of the inhibitory killer cell lectin-like receptor G1 (KLRG1) expressing cellular subset. Furthermore, higher plasma levels of the natural KLRG1 ligand E-cadherin were detected in CLL patients compared to normal donor controls. The predominance of KLRG1+ expression within CD8+ T cells in conjunction with increased systemic soluble E-cadherin might significantly contribute to CLL immune dysfunction and might additionally represent an important component of the CLL microenvironment. 相似文献
Sevoflurane has been demonstrated to vasodilate the foeto-placental vasculature. We aimed to determine the contribution of modulation of potassium and calcium channel function to the vasodilatory effect of sevoflurane in isolated human chorionic plate arterial rings.
Methods
Quadruplicate ex vivo human chorionic plate arterial rings were used in all studies. Series 1 and 2 examined the role of the K+ channel in sevoflurane-mediated vasodilation. Separate experiments examined whether tetraethylammonium, which blocks large conductance calcium activated K+ (KCa++) channels (Series 1A+B) or glibenclamide, which blocks the ATP sensitive K+ (KATP) channel (Series 2), modulated sevoflurane-mediated vasodilation. Series 3 – 5 examined the role of the Ca++ channel in sevoflurane induced vasodilation. Separate experiments examined whether verapamil, which blocks the sarcolemmal voltage-operated Ca++ channel (Series 3), SK&F 96365 an inhibitor of sarcolemmal voltage-independent Ca++ channels (Series 4A+B), or ryanodine an inhibitor of the sarcoplasmic reticulum Ca++ channel (Series 5A+B), modulated sevoflurane-mediated vasodilation.
Results
Sevoflurane produced dose dependent vasodilatation of chorionic plate arterial rings in all studies. Prior blockade of the KCa++ and KATP channels augmented the vasodilator effects of sevoflurane. Furthermore, exposure of rings to sevoflurane in advance of TEA occluded the effects of TEA. Taken together, these findings suggest that sevoflurane blocks K+ channels. Blockade of the voltage-operated Ca++channels inhibited the vasodilator effects of sevoflurane. In contrast, blockade of the voltage-independent and sarcoplasmic reticulum Ca++channels did not alter sevoflurane vasodilation.
Conclusion
Sevoflurane appears to block chorionic arterial KCa++ and KATP channels. Sevoflurane also blocks voltage-operated calcium channels, and exerts a net vasodilatory effect in the in vitro foeto-placental circulation. 相似文献
Conformations of three pairs of dehydropeptides with the opposite configuration of the ΔPhe residue, Boc-Gly-ΔZ/EPhe-Phe-p-NA (Z-p-NA and E-p-NA), Boc-Gly-ΔZ/EPhe-Phe-OMe (Z-OMe and E-OMe), and Boc-Gly-ΔZ/EPhe-Phe-OH (Z-OH and E-OH) were compared on the basis of CD and NMR studies in MeOH, TFE, and DMSO. The CD results were used as the additional input data for the NMR-based calculations of the detailed solution conformations of the peptides. It was found that Z-p-NA, E-p-NA, Z-OMe, and Z-OH adopt the β-turn conformations and E-OMe and E-OH are unordered. There are two overlapping type III β-turns in Z-p-NA, type II’ β-turn in E-p-NA, and type II β-turn in Z-OMe and Z-OH. The results obtained indicate that in the case of methyl esters and peptides with a free carboxyl group, ΔZPhe is a much stronger inducer of ordered conformations than ΔEPhe. It was also found that temperature coefficients of the amide protons are not reliable indicators of intramolecular hydrogen bonds donors in small peptides. 相似文献
The mechanism of phenanthridines synthesis from the nitrogenation of 2-acetylbiphenyls (1) by TMSN3 in TFA has been studied by DFT calculations. Results at the B3LYP/6-311G(d) level showed that: 1) reaction of TMSN3/HN3 with the protonated form of 1 (1H+), which generates the key intermediate Cx+ by removal of TMSOH/H2O, is the rate determining step, and TMSN3 as the nitrogen source is certainly preferred over HN3. 2) from Cx+, the two pathways leading to 2xH+ and 3xH+ are both thermodynamically and kinetically feasible and competitive to each other. 3) The high barriers of the reverse reactions suggest that the ratio of the final products 2x:3x is determined by the branching ratio of reaction rates of Cx+ to intermediates Dx+ in pass_I and Ex+ in pass_II.
Graphical Abstract DFT results indicate that the replacement of -OH by -N3 which generates Cx+ controls the consumption rate of 1x H+, and the ratio of Cx+ transforms to Dx+ and Cx+ transforms to Ex+ (k:k') determines the final ratio of products 2x:3x.
This study established an efficient method of regenerating plants ofFicus lyrataand producing purple-leavedF. lyrataplants through genetic transformation using aVvMybA1gene of grapevine.
Abstract
Ficus lyrata, a species with unique violin- or guitar-shaped leaves, was regenerated from leaf-derived calli cultured on Murashige and Skoog (MS) basal medium supplemented with 4.5 μM N-phenyl-N’-1, 2, 3-thiadiazol-5-yl urea (TDZ) and 0.5 μM α-naphthalene acetic acid (NAA). Leaf discs were inoculated with Agrobacterium tumefaciens strain EHA 105 harboring a binary vector DEAT that contains the VvMybA1 gene and neomycin phosphotransferase (npt II) gene and subsequently cultured on the established regeneration medium supplemented with 100 mg l?1 kanamycin. Results showed that 87.5 % of the leaf discs produced kanamycin-resistant callus, and 68.8 % of them produced adventitious shoots. Transgenic plants with three leaf colors including green, green-purple, and purple were produced. Regular and quantitative real-time PCR analyses confirmed the integration of transgenes into the host genome. Semi-quantitative RT-PCR analysis indicated that the VvMybA1 gene was responsible for the purple-colored phenotype. Purple-leaved plants with strong color stability grew vigorously in a greenhouse. This study illustrated the feasibility of using a genetically engineered VvMybA1 gene for drastic modification of leaf color of an important woody ornamental plant. 相似文献
N-(2-Benzothiazolyl)- and N-(6-methoxy-2-benzothiazolyl)cyanoacetamides 4, 5 resulted in the reaction of 2-aminobenzothiazole 1 or its 6-methoxy derivative 2 with 1-cyanoacetyl-3,5-dimethylpyrazole 3. Both cyanoacetylamides 4 and 5 have been transformed into the corresponding 2-oxo-2H-pyrimido[2,1-b]-benzothiazole-3-carbonitrile 8 and its 8-methoxy derivative 9 by reaction with triethyl orthoformate, followed by cyclization. 相似文献
Novel 1,6-diaryl-5,7(1H)dioxo(dithio)-2,3-dihydroimidazo[1,2-a][1, 3, 5]triazines 8, and 9 were synthesized by cyclization of the respective 1-(imidazolin-2-yl)ureas 4 or thioureas 6 with phosgene or thiophosgene in the presence of bases. 1-Aryl-2-aminoimidazolines 1 reacting with arylisocyanates 2 or arylisothiocyanates 3 form a mixture of isomeric imidazolin-2-yl 4 and 6 and imidazolin-3-yl 5 and 7 urea or thiourea derivatives. Isomers 4 and 6 can be easily separated and used for the cyclization reaction. The structures of the main intermediates and the final target compounds were confirmed by 1H-NMR spectral analysis. Discussion of the possible course of the reactions is also presented. 相似文献
For the first time the putativeNSP2gene in chickpea has been identified using pairs of NILs differing for theRn1/rn1nodulation gene that was located in LG5 of chickpea genetic map.
Abstract
An intraspecific cross between the mutant non-nodulating genotype PM233, carrying the recessive gene rn1, and the wild-type CA2139 was used to develop two pairs of near-isogenic lines (NILs) for nodulation in chickpea. These pairs of NILs were characterized using sequence tagged microsatellite site (STMS) markers distributed across different linkage groups (LGs) of the chickpea genetic map leading to the detection of polymorphic markers located in LG5. Using this information, together with the genome annotation in Medicago truncatula, a candidate gene (NSP2) known to be involved in nodulation pathway was selected for mapping in chickpea. The full length sequence obtained in chickpea wild-type (CaNSP2) was 1,503 bp. Linkage analysis in an F3 population of 118 plants derived from the cross between the pair of NILS NIL7-2A (nod) × NIL7-2B (non-nod) revealed a co-localization between CaNSP2 and Rn1 gene. These data implicate the CaNSP2 gene as a candidate for identity to Rn1, and suggest that it could act in the nodulation signaling transduction pathway similarly to that in other legumes species. 相似文献
The gas phase molecular structure of a single isolated molecule of [Ag(Etnic)2NO3];1 where Etnic = Ethylnicotinate was calculated using B3LYP method. The H-bonding interaction between 1 with one (complex 2) and two (complex 3) water molecules together with the dimeric formula [Ag(Etnic)2NO3]2;4 and the tetrameric formula [Ag(Etnic)2NO3]4;5 were calculated using the same level of theory to model the effect of intermolecular interactions and molecular packing on the molecular structure of the titled complex. The H-bond dissociation energies of complexes 2 and 3 were calculated to be in the range of 12.220–14.253 and 30.106–31.055 kcal?mol?1, respectively, indicating the formation of relatively strong H-bonds between 1 and water molecules. The calculations predict bidentate nitrate ligand in the case of 1 and 2, leading to distorted tetrahedral geometry around the silver ion with longer Ag–O distances in case of 2 compared to 1, while 3 has a unidentate nitrate ligand leading to a distorted trigonal planar geometry. The packing of two [Ag(Etnic)2NO3] complex units; 4 does not affect the molecular geometry around Ag(I) ion compared to 1. In the case of 5, the two asymmetric units of the formula [Ag(Etnic)2NO3] differ in the bonding mode of the nitrate group, where the geometry around the silver ion is distorted tetrahedral in one unit and trigonal planar in the other. The calculations predicted almost no change in the charge densities at the different atomic sites except at the sites involved in the C–H?O interactions as well as at the coordinated nitrogen of the pyridine ring.
Figure
Molecular structure (left) and electrostatic potentials mapped on the electron density surface (right) calculated by DFT/B3LYP method for Etnic, and complexes 1 and 2相似文献
Cyclometalated RuII derivatives of 2-phenylpyridine (Hphpy) [Ru(phpy)(bpy)2]Cl (1a) and [Ru(phpy)(phen)2]Cl (1b) (bpy is 2,2′-bipyridine, phen is 1,10-phenanthroline) behave as noncompetitive inhibitors of glucose oxidase from Aspergillus niger in the enzyme-catalyzed oxidation of d-glucose by O2 into the corresponding lactone at pH 5.0 and 25 °C. The enzymatic activity has been measured by monitoring the O2 consumption. The inhibition constants Ki are 0.036 and 0.017 M for 1a and 1b, respectively, indicating that 1b inhibits the enzymatic activity more efficiently than 1a. The well-known coordination compound [Ru(bpy)3]Cl2 (2) behaves, in contrast, as a competitive inhibitor, with Ki = 0.018 M under the same conditions. The monophasic consumption of O2 in the case of 1a, 1b, and 2 is replaced by a distinct two-phase kinetics in the presence of the cyclometalated RuIII compound [Ru(phpy)(bpy)2]Cl2 (3), which was obtained from 1a in the presence of a large excess of H2O2 and the iron TAML activator. Interestingly, the rates of the first and the second phases are influenced by 3 in a different way. The rate of the first phase is noticeably higher in the presence of RuIII, although the dependence is nonmonotonic and maximal acceleration is observed at the lowest loadings of 3. The rate of the second phase decreases monotonically on increasing the concentration of the ruthenium complex in solution. The nonmonotonic action of 3 was confirmed by using the doubly cyclometalated RuIII derivative [Ru(phpy)2(bpy)]Cl. The diverse rate variations induced by 3 accounted for acceleration by RuIII of the O2 reduction by the reduced form of glucose oxidase during the first phase, which ceases after the enzymatic reduction of RuIII to the RuII species, the latter behaving similarly to 1a as the inhibitor of the enzyme. 相似文献
Density functional theory (DFT) with relativistic corrections of zero-order regular approximation (ZORA) has been applied to explore the reaction mechanisms of ethane dehydrogenation by Zr atom with triplet and singlet spin-states. Among the complicated minimum energy reaction path, the available states involves three transition states (TS), and four stationary states (1) to (4) and one intersystem crossing with spin-flip (marked by ?): 3Zr + C2H6 → 3Zr-CH3-CH3 (31) → 3TS1/2 → 3ZrH-CH2-CH3 (32) → 3TS2/3 ? 1ZrH2-CH2 = CH2 (13) → 1TS3/4 → 1ZrH3-CH = CH2 (14). The minimum energy crossing point is determined with the help of the DFT fractional-occupation-number (FON) approach. The spin inversion leads the reaction pathway transferring from the triplet potential energy surface (PES) to the singlet’s accompanying with the activation of the second C-H bond. The overall reaction is calculated to be exothermic by about 231 kJ mol?1. Frequency and NBO analysis are also applied to confirm with the experimental observed data.
Reaction 3 Zr + C 2 H 6 → 3 ZrH ? CH 2 ? CH 3 ? 1 ZrH 2 ? CH 2 = CH 2 → 1 ZrH 3 ? CH = CH 2 $ {}^{\mathbf{3}}\mathrm{Zr}+{\mathrm{C}}_{\mathbf{2}}{\mathrm{H}}_{\mathbf{6}}{\to}^{\mathbf{3}}\mathrm{Zr}\mathrm{H}-{\mathrm{C}\mathrm{H}}_{\mathbf{2}}-{\mathrm{C}\mathrm{H}}_{\mathbf{3}}{\Rightarrow}^{\mathbf{1}}{\mathrm{ZrH}}_2-{\mathrm{C}\mathrm{H}}_2={\mathrm{C}\mathrm{H}}_2{\to}^{\mathbf{1}}{\mathrm{ZrH}}_{\mathbf{3}}-\mathrm{CH}={\mathrm{C}\mathrm{H}}_{\mathbf{2}} $proceeds via spin-flip surface hopping over several transition states has been investigated. The minimum energy crossing point is determined with the help of the DFT fractional-occupation-number (FON) approach. 相似文献