Activity

Enzymatic reactions consist of several steps (i) a weak binding event of the substrate to the enzyme, (ii) an induced fit or a protein conformational transition upon ligand binding, (iii) the chemical reaction, and (iv) the release of the product. Here we focus on step iii of the reaction of a DNA polymerase, HIV RT, with a nucleotide. We determine the rate and the free energy profile for the addition of a nucleotide to a DNA strand using a combination of a QM/MM model, the string method, and exact Milestoning. The barrier height and the time scale of the reaction are consistent with experiment. We show that the observables (free energies and mean first passage time) converge rapidly, as a function of the Milestoning iteration number. We also consider the substitution of an oxygen of the incoming nucleotide by a nonbridging sulfur atom and its impact on the enzymatic reaction. This substitution has been suggested in the past as a tool to examine the influence of the chemical step on the overall rate. Our joint computational and experimental study suggests that the impact of the substitution is small. Computationally, the differences between the two are within the estimated error bars. Experiments suggest a small difference. Finally, we examine step i, the weak binding of the nucleotide to the protein surface. We suggest that this step has only a small contribution to the selectivity of the enzyme. Comments are made on the impact of these steps on the overall mechanism.Three new caryophyllane-type sesquiterpenoids, linariophyllenes A-C (1-3), two new hamamelitol derivatives, linaritols A (4) and B (5), two new chromones, linariosides A (6) and B (7), and three known chromones, cnidimol C (8), monnieriside A (9), and undulatoside A (10), were identified from the aerial parts of Evolvulus linarioides. The structures of these compounds were elucidated by NMR, MS, and IR data. The absolute configurations of compounds 1-5 and 7 were established via electronic circular dichroism data. see more The anti-inflammatory potential of compounds 1-5 and 7-10 was evaluated by determining their ability to inhibit the production of nitric oxide (NO) and proinflammatory cytokine IL-1β by stimulated J774 macrophages. Compounds tested at noncytotoxic concentrations inhibited NO production by macrophages, exhibiting IC50 values between 17.8 and 66.2 μM, and inhibited IL-1β production by stimulated macrophages by 72.7-96.2%.A weakly coordinating, carbonyl-assisted C-H activation of aromatic systems with α,β-unsaturated ketone and subsequent aldol condensation has been developed using a Cp*Co(CO)I2 catalyst. The developed method is the first example of indene synthesis by cobalt-catalyzed C-H activation. In addition, the reaction requires mild reaction conditions and easily accessible starting materials, and it shows excellent functional group compatibility.In the prodrug research field, information obtained from traditional end point biochemical assays in drug effect studies could provide neither the dynamic processes nor heterogeneous responses of individual cells. In situ imaging microscopy techniques, especially fluorescence lifetime imaging microscopy (FLIM), could fulfill these requirements. In this work, we used FLIM techniques to observe the entry and release of doxorubicin (Dox)-Cu complexes in live KYSE150 cells. The Dox-Cu complex has weaker fluorescence signals but similar lifetime values as compared to the raw Dox, whose fluorescence could be released by the addition of biothiol compound (such as glutathione). The cell viability results indicated that the Dox-Cu compound has a satisfactory killing effect on KYSE150 cells. The FLIM data showed that free doxorubicin was released from Dox-Cu complexes in cytoplasm of KYSE150 cells and then accumulated in the nucleus. After 90 min administration, the fluorescence lifetime signals reached 1.21 and 1.46 ns in the cytoplasm and nucleus, respectively, reflecting the transformation and transportation of Dox-Cu complexes. In conclusion, this work provides a satisfactory example for the research of prodrug monitored by FLIM techniques, expanding the useful applications of FLIM technique in drug development.The ethanolic root extract of Berberis brevissima afforded a new bisbenzylisoquinoline alkaloid, 13-nitrochondrofoline (2), and two known bisbenzylisoquinoline alkaloids, chondrofoline (1) and curine (4). The acetylation of chondrofoline (1) gave O-acetylchondrofoline (3). The dimeric structures of 1 and 2 were studied through variable-temperature 1H NMR spectroscopy at 25, 40, 60, and 80 °C and conformational analysis, using density functional theory employing the M06-2X functional and the 6-31G* basis set. The in vitro antitrypanosomal activity of compounds 1, 2, 3, and 4 against Trypanosoma brucei showed significant potential with MIC values of 2.6, 2.2, 2.3, and 3.8 μM, respectively. Molecular docking evaluation of alkaloids 1, 2, 3, and 4 against known T. brucei protein targets revealed T. brucei phosphodiesterase B1 to be the preferred target. The docking energies of the alkaloids with Tb6PGL (PDB 3EB9) ranged from -88.8 to -106.0 kJ/mol and was comparable to the cocrystallized ligand, citrate (Edock = -78.3 kJ/mol). It seems reasonable that the curine alkaloids may compete with the natural substrates for these protein targets and serve as leads in designing and developing more potent and selective drugs against T. brucei.The cyclic dimeric peptide 1229U91 (GR231118) has an unusual structure and displays potent, insurmountable antagonism of the Y1 receptor. To probe the structural basis for this activity, we have prepared ring size variants and heterodimeric compounds, identifying the specific residues underpinning the mechanism of 1229U91 binding. The homodimeric structure was shown to be dispensible, with analogues lacking key pharmacophoric residues in one dimer arm retaining high antagonist affinity. Compounds 11d-h also showed enhanced Y1R selectivity over Y4R compared to 1229U91.Direct dynamics trajectory simulations were carried out for the NO2 oxidation of 1-ethyl-3-methylimidazolium dicyanamide (EMIM+DCA-), which were aimed at probing the nature of the primary and secondary reactions in the system. Guided by trajectory results, reaction coordinates and potential energy diagrams were mapped out for NO2 with EMIM+DCA-, as well as with its analogues 1-butyl-3-methylimidazolium dicyanamide (BMIM+DCA-) and 1-allyl-3-methylimidazolium dicyanamide (AMIM+DCA-). Reactions of the dialkylimidazolium-dicyanamide (DCA) ionic liquids (ILs) are all initiated by proton transfer and/or alkyl abstraction between 1,3-dialkylimidazolium cations and DCA- anion, of which two exoergic pathways are particularly relevant to their oxidation activities. One pathway is the transfer of a Hβ-proton from the ethyl, butyl, or allyl group of the dialkylimidazolium cation to DCA- that results in the concomitant elimination of the corresponding alkyl as a neutral alkene, and the other pathway is the alkyl abstraction by DCA- via a second order nucleophilic substitution (SN2) mechanism.