A study of the interactions among adenosine triphosphate, epinephrine, and magnesium ions

The interactions among adenosine triphosphate, Mg⁺², and epinephrine at pH's below 7.0 have been studied by observing the effects of these interactions on the chemical shifts and line widths of their ¹H and ³¹P nuclear magnetic resonance spectra. Mg⁺² is tightly bound by the β- and γ-phosphate groups of adenosine triphosphate and there is a weak association between this chelate and epinephrine. In the ternary complex, the aromatic ring of epinephrine overlaps the purine ring of adenosine triphosphate and there appears to be an ionic interaction between the protonated amino group and the α-phosphate of adenosine triphosphate. It was also found that dichloroisoproterenol forms essentially the same type of ternary complex.     

DNA Minor Groove Induced Dimerization of Heterocyclic Cations: Compound Structure, Binding Affinity, and Specificity for a TTAA Site

With the increasing number and variations of genome sequences available, control of gene expression with synthetic, cell-permeable molecules is within reach. The variety of sequence-specific binding agents is, however, still quite limited. Many minor groove binding agents selectivity recognize AT over GC sequences but have less ability to distinguish among different AT sequences. The goal with this article is to develop compounds that can bind selectively to different AT sequences. A number of studies indicate that AATT and TTAA sequences have significantly different physical and interaction properties and different requirements for minor groove recognition. Although it has been difficult to get minor groove binding at TTAA, DB293, a phenyl-furan-benzimidazole diamidine, was found to bind as a strong, cooperative dimer at TTAA but with no selectivity over AATT. In order to improve selectivity, we made modifications to each unit of DB293. Binding affinities and stoichiometries obtained from biosensor-surface plasmon resonance experiments show that DB1003, a furan-furan-benzimidazole diamidine, binds strongly to TTAA as a dimer and has selectivity (K_TTAA/K_AATT = 6). CD and DNase I footprinting studies confirmed the preference of this compound for TTAA. In summary, (i) a favorable stacking surface provided by the pi system, (ii) H-bond donors to interact with TA base pairs at the floor of the groove provided by a benzimidazole (or indole) -NH and amidines, and (iii) appropriate curvature of the dimer complex to match the curvature of the minor groove play important roles in differentiating the TTAA and AATT minor grooves.     

Structure-based design,synthesis, and binding mode analysis of novel and potent chymase inhibitors

Based on insight from the X-ray crystal structure of human chymase in complex with compound 1, a lactam carbonyl of the diazepane core was exchanged with O-substituted oxyimino group, leading to amidoxime derivatives. This modification resulted in highly potent chymase inhibitors, such as O-phenylamidoxime 5f. X-ray crystal structure analysis indicated that compound 5f induced movement of the Leu99 and Tyr94 side chains at the S2 site, and the increase in inhibitory activity of O-phenyl amidoxime derivatives suggested that the O-phenyl moiety interacted with the Tyr94 residue. Surface plasmon resonance experiments showed that compound 5f had slower association and dissociation kinetics and the calculated residence time of compound 5f to human chymase was extended compared to that of amide compound 1.     

Carotenoid radical cations and dications: EPR, optical, and electrochemical studies

Investigations of the structure and properties of paramagnetic carotenoid radical cations and diamagnetic carotenoid dications using electron paramagnetic resonance (EPR) and electron-nuclear double resonance (ENDOR) spectroscopy in conjunction with electrochemical, optical, and HPLC measurements, and molecular orbital calculations are described. These methods were applied to determine how the carotenoid radical cations and dications can be formed, their electron-transfer properties and stability in various media, and the mechanism by which carotenoid radical cations can isomerize.     

The effect of short-term fasting on liver and skeletal muscle lipid, glucose, and energy metabolism in healthy women and men

Fasting promotes triglyceride (TG) accumulation in lean tissues of some animals, but the effect in humans is unknown. Additionally, fasting lipolysis is sexually dimorphic in humans, suggesting that lean tissue TG accumulation and metabolism may differ between women and men. This study investigated lean tissue TG content and metabolism in women and men during extended fasting. Liver and muscle TG content were measured by magnetic resonance spectroscopy during a 48-h fast in healthy men and women. Whole-body and hepatic carbohydrate, lipid, and energy metabolism were also evaluated using biochemical, calorimetric, and stable isotope tracer techniques. As expected, postabsorptive plasma fatty acids (FAs) were higher in women than in men but increased more rapidly in men with the onset of early starvation. Concurrently, sexual dimorphism was apparent in lean tissue TG accumulation during the fast, occurring in livers of men but in muscles of women. Despite differences in lean tissue TG distribution, men and women had identical fasting responses in whole-body and hepatic glucose and oxidative metabolism. In conclusion, TG accumulated in livers of men but in muscles of women during extended fasting. This sexual dimorphism was related to differential fasting plasma FA concentrations but not to whole body or hepatic utilization of this substrate.     

1H, 13C, and 15N NMR assignments of the actinoporin Sticholysin I

Sticholysin I is an actinoporin, a pore forming toxin, of 176 aminoacids produced by the sea anemone Stichodactyla heliantus. Isotopically labelled ¹³C/¹⁵N recombinant protein was produced in E. coli. Here we report the complete NMR ¹⁵N, ¹³C and ¹H chemical shifts assignments of Stn I at pH 4.0 and 25°C (BMRB No. 15927).     

Solubilization, stabilization, and purification of chemokine receptors using biosensor technology

Establishing solubilization conditions for membrane-associated receptors is often a tedious empirical process. Here we describe a novel application of SPR biosensor technology to screen solubilization conditions automatically and to assess receptor activity directly. We focus on two chemokine receptors, CXCR4 and CCR5, which are important in HIV cell invasion. The autosampler in Biacore 3000 permitted whole cells expressing C-terminally tagged receptors to be automatically lysed under a given solubilization condition and the lysates to be injected over an antibody surface. The total amount of solubilized receptor could be quantitated from the antibody capture level, whereas the amount of active receptor could be quantitated using a subsequent injection of conformationally sensitive antibody or protein. Using this approach, we identified detergent/lipid/buffer combinations that enhanced and maintained receptor activity. We also used the biosensor to demonstrate CD4-dependent binding of gp120 to solubilized CCR5 and to develop affinity chromatography-based purification methods that increased receptor activity more than 300%. Together, these results illustrate the benefits of using the biosensor as a tool for isolating functional membrane receptors and for analyzing ligand/receptor interactions.     

Preparation, characterization and in vivo assessment of Gd-albumin and Gd-dendrimer conjugates as intravascular contrast-enhancing agents for MRI

We report in vivo and in vitro MRI properties of six gadolinium-dendrimer and gadolinium-albumin conjugates of derivatized acyclic diethylenetriamine-N,N′,N′,N″, N″-pentaacetic acid (1B4M) and macrocyclic 1,4,7,10-tetraazacyclododecane-N,N′,N″,N?-tetraacetic acid (C-DOTA). The three albumin-based agents have comparable protein to chelate ratios (1:16-18) as well as molar relaxivity (8.8-10.4 mM^− 1 s^− 1). The three dendrimer based agents have blood clearance half-lives ranging from 17 to 66 min while that of the three albumin-based agents are comparable to one another (40-47 min). The dynamic image obtained from use of the albumin conjugate based on the macrocycle (C-DOTA) showed a higher contrast compared to the remaining two albumin based agents. Our conclusion from all of the results is that the macrocyclic-based (DOTA) agents are more suitable than the acyclic-based (1B4M) agent for in vivo use based on their MRI properties combined with the kinetic inertness property associated with the more stable Gd(III) DOTA complex.     

Sequence-specific 1H, 13C, and 15N backbone assignment of the activated 21 kDa GTPase rRheb

Ras homologue enriched in brain (Rheb) is a small GTPase that plays an important role in tuberous sclerosis. Here we present the backbone assignments of activated rRheb in complex with the non-hydrolisable GTP analogue GppNHp. These assignments now provide a basis for the analysis of rRheb’s interaction with putative effectors in order to further elucidate the physiological function of this GTPase and its role in the regulation of neuronal cell volume as well as in tuberous sclerosis.     

Backbone and side-chain 1H, 13C, 15N resonance assignments of rat lipocalin2

Lipocalin2 plays an important role in the innate immune system. In this article we report the backbone and side-chain resonance assignments of rat lipocalin2 (rLcn2). These assignments provide a basis for determining the structure and dynamics of rLcn2. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

寄生隐丛赤壳菌致病毒素Cp-I的分离纯化和结构分析

寄生隐丛赤壳菌Cryphonectria parasitica菌株经液体培养,石油醚萃取其发酵液获得对板栗带叶嫩枝具有致萎活性的粗提物,以氯仿:石油醚:甲醇(6:2:2)作洗脱剂,粗提物经硅胶色谱分离,共得到3组纯组份,其中第1组份(Cp-I)对板栗幼苗致萎活性较高。质谱、核磁共振和红外光谱测定表明Cp-I分子量为278,化学式为C16H22O4。