Nonequilibrium capillary electrophoresis of equilibrium mixtures--a single experiment reveals equilibrium and kinetic parameters of protein-DNA interactions

We introduce a novel electrophoretic method, nonequilibrium capillary electrophoresis of equilibrium mixtures (NECEEM), and demonstrate its use for studying protein-DNA interactions. The equilibrium mixture of protein and DNA contains three components: free protein, free DNA, and the protein-DNA complex. A short plug of such a mixture is injected into the capillary, and the three components are separated under nonequilibrium conditions. The resulting electropherograms are composed of characteristic peaks and exponential curves. An easy nonnumerical analysis of a single electropherogram reveals two parameters: the equilibrium binding constant and the monomolecular rate constant of complex decay. The bimolecular rate constant of complex formation can then be calculated as the product of the two experimentally determined constants. NECEEM was applied to study the interaction between single-stranded DNA binding protein and a fluorescently labeled 15-mer oligonucleotide. It allowed us to measure for the first time the rate constant of complex decay for this important protein-DNA pair, k-1 = 0.03 s-1. The value of the equilibrium binding constant, Kb = 3.6 x 10-6 M-1, was in good agreement with those measured by other methods. As low as 10-18 mol of the protein was sufficient for the measurements. Thus, the new method is simple, informative, and highly sensitive. Moreover, it can be equally applied to other noncovalent protein-ligand complexes. These features of NECEEM make this method an indispensable tool in studies of macromolecular interactions. They also emphasize the potential role of NECEEM in the development of extremely sensitive protein assays using nucleotide aptamers.     

Accurate and efficient generalized born model based on solvent accessibility: derivation and application for LogP octanol/water prediction and flexible peptide docking

A novel method for fast and accurate evaluation of the generalized Born radii in macromolecular solvation electrostatics calculations is proposed, based on the solvent accessibility of the first two solvation layers around an atom. The reverse generalized Born radii calculated by the method have correlation coefficient of 98.7% and RMSD of 0.031 A(-1) with the values obtained using a precise but significantly slower numerical boundary element solution. The method is applied to derive an estimate of the free solvation energy difference between octanol and water and to predict LogP octanol-water. A nine-parameter model is optimized on an 81 compound training set and applied to predict LogP(ow) for an external evaluation set of 19 drug molecules with RMSD of 0.9. The new GB approximation is also tested in Monte Carlo docking simulations of the fully flexible p53 peptide fragment to MDM2. The best energy solution found in the simulations has RMSD of 2.8 A to the X-ray structure.     

有水条件下路易斯酸B(C6F5)3对醛酮的选择性催化还原

近年路易斯酸B（C₆F₅）₃催化的醛酮还原反应研究表明，路易斯酸B（C₆F₅）₃也可以作为一种"耐水"的催化剂在"有水"条件下进行催化反应.这些研究成果对进一步扩展受限路易斯酸碱对（FLPs）化学在水相中的应用及其发展至关重要.以硅烷作为还原剂，在路易斯酸B（C₆F₅）₃催化下成功对14种不同取代的醛酮在温和条件下实现了高效地还原成醇反应，产率可高达100%.通过核磁共振分析对反应机理进行的研究表明，在有水条件下，底物羰基氧与催化剂硼烷之间存在以水介导的相互作用，即可能存在"R¹R²C=O--H-O（-H）--B（C₆F₅）₃"的三组分络合形式.对不同路易斯碱与水及硼烷所组成的"LB--H-O（-H）--B（C₆F₅）₃"三组分复合物之间对应原子间距进行了比较研究，发现这种以水介导的相互作用对醛酮的羰基不仅具有质子化的活化作用，而且与硼烷的络合作用也更有利于硅烷作为还原剂在有水条件下使用.所报道的以硅烷作为还原剂，B（C₆F₅）₃催化的醛酮直接还原成醇反应，首次实现了FLPs在真正有水条件下的催化反应，并更进一步证明了在以硅烷作为还原剂时，体系中存在的水不仅会参与催化反应并会促使反应的进行.对有水条件下FLPs催化的醛酮还原的选择性及其催化反应机理，以及对其他反应的影响还需要更深入的研究.     

Aqueous solution chemistry of [Mo3CuSe4]n+ (n = 4, 5) and [W3CuQ4]5+ (Q = S, Se) clusters

The triangular cluster [Mo3Se4(H2O)9]4+ reacts with Cu turnings to give a new heterometallic cuboidal cluster [Mo3CuSe4(H2O)10]4+(purple; UV/Vis lambda(epsilon): 352(3907), 509(2613)). The reaction of [Mo3Se4(H2O)9]4+ with CuCl afforded the 5+ cube [Mo3CuSe4(H2O)10]5+(red; UV/Vis lambda(epsilon): 356(5406), 500(3477)). In contrast, [W3Se4(H2O)9]4+ both with Cu and CuCl gives the 5+ cube, [W3CuSe4(H2O)10]5+(yellow-green; UV/Vis lambda(epsilon): 312(5327), 419(3256) and 628(680)). Cyclic voltammetry of [M3CuQ4(H2O)10]5+ in 2 M HCl (M = Mo, W; Q = S, Se) shows a reversible one-electron reduction wave for the Mo clusters, but no reduction occurs for the W clusters prior to H+ reduction. In HCl solutions, Cl is coordinated to the Cu site of the clusters, alongside some less extensive coordination to Mo and W, and for [W3(CuCl)S4(H2O)6Cl3]+, isolated as the supramolecular adduct with cucurbit[6]uril, [W3(CuCl)S4(H2O)6Cl3]2Cl2 x C36H36N24O12 x 12H2O, the crystal structure was determined (Cu-W 2.856(4) angstroms, W-W 2.7432(15) angstroms, Cu-Cl 2.167(13) angstroms).     

Molecular dynamics simulation of the ligand binding domain of mGluR1 in response to agonist and antagonist binding

The interdomain movements of the ligand binding domain (LBD) of mGluR1 in response to agonist or antagonist binding are studied by 2 ns molecular dynamics (MD) simulations. Our results indicate that MD is able to reproduce many of the experimentally determined features of the open and closed conformations of LBD. Analysis of the ligand behavior over time allows to delineate some of the molecular determinants responsible for the agonist-induced or antagonist-blocked LBD responses.     

5,6-Bis(dimethylamino)acenaphthylene as an activated alkene and ‘proton sponge’ in halogenation reactions

It has been shown that 5,6-bis(dimethylamino)acenaphthylene in its reactions with X₂ (X=Cl, Br, I) and N-X-succinimides behaves simultaneously as an electron-rich alkene or arene and proton sponge. Thus, addition of bromine or iodine to the C(1)C(2) bond is followed by immediate dehydrohalogenation leading to the formation of the corresponding 1(2)-(di)halogenoacenaphthylenes in good yields. Reaction with chlorine enables isolation of only 1,4,7-trichloro-5,6-bis(dimethylamino)acenaphthylene. With N-halosuccinimides, the halogenation is directed mainly by the steric bulk of the entering halogen and then by solvent polarity thus allowing the regioselective preparation of 1(2)- or 4(7)-(di)halides. Introduction of the third and fourth bromine atoms, but not chlorine, is accomplished by mono-N-demethylation. The pK_a values of some new derivatives of acenaphthene and acenaphthylene proton sponges were measured by competitive transprotonation ¹H NMR spectroscopy technique in DMSO. The X-ray molecular structures of 4,7-dichloro-5,6-bis(dimethylamino)acenaphthylene and its monoprotonated form are reported.     

Template copolymerization near a patterned surface: computer simulation

We perform a Monte Carlo simulation of irreversible template copolymerization near a chemically heterogeneous surface with a regular distribution of discrete adsorption sites that selectively adsorb from solution one of the two polymerizing monomers and the corresponding chain segments. In the polymerization model, the chain propagation process is simulated by adding individual monomers to the end of growing macroradical. We focus in this paper on the influence of polymerization rate, adsorption energy, and the distance between adsorption sites on the chain conformation and the primary sequence of the resulting two-letter (AB) copolymers and, specifically, on the coupling between polymerization and adsorption. The conditions for the realization of conformation-dependent copolymerization are formulated. For this regime, we observe the formation of a quasiregular copolymer with two types of alternating sections. One of them contains randomly distributed A and B segments. The second one consists mainly of strongly adsorbed A segments. It is found that the average length of the random sections is proportional to the distance between the nearest neighbor adsorption sites. The average length of the A-rich sections is determined by the "adsorption capacity" of adsorption site. By varying the strength of the effective monomer-substrate interaction and the distribution of adsorption sites on the substrate, the copolymers with different surface-induced primary sequences can be designed and synthesized in a controlled fashion.     

Mono- and bis-[2-(P,P-dimethylphosphanyl)ethyl]tetramethylcyclopentadienyl zirconium(IV) complexes: Synthesis and structural studies in crystalline state and in solutions

Synthetic routines for a new ligand C₅Me₄CH₂CH₂PMe₂ (2b) in forms of its Li- (2b-Li), Na- (2b-Na) salts and in the CH-form (2b-H), as well as for silanes Me₃Si-C₅H₄CH₂CH₂PMe₂ (3a) and Me₃Si-C₅Me₄CH₂CH₂PMe₂ (3b) have been developed. On the basis of it, new half-sandwich [η⁵:η¹-κP-C₅H₄CH₂CH₂PMe₂]ZrCl₃ (4a), [η⁵:η¹-κP-C₅Me₄CH₂CH₂PMe₂]ZrCl₃ (4b) and sandwich [η⁵-C₅Me₄CH₂CH₂PMe₂]₂ZrCl₂ (5), [η⁵-C₅Me₄CH₂CH₂PMe₂][η⁵-C₅Me₅]ZrCl₃ (6) complexes of Zr(IV) have been prepared and characterized. Along with them, the first example of X-ray structurally characterized dinuclear Zr(IV) complex incorporating both sandwich (6) and half-sandwich (4b) moieties linked one to another by means of Zr ← P coordination bond 7, has been described. Formation of an analogously organized trinuclear complex 8, built from one sandwich fragment of 5 and two half-sandwich fragments of 4b was proved by NMR spectroscopy methods. Molecular structures of half-sandwich complexes in their solvent-free dimeric forms (4a and 4b) and as 1:1 adducts with THF (4a-THF and 4b-THF) along with those of dinuclear complex 7 have been established by X-ray diffraction analyses. The dynamic behavior for di- and trinuclear complexes 7 and 8, due to the intermolecular dissociation-coordination of the Me₂P-groups in THF-d₈ solutions has been studied by variable-temperature NMR spectroscopy.     

Oxygen vacancy formation and ionic transport in Sr4Fe6O13± δ

Oxygen permeation through dense Sr₄Fe₆O_{13± δ} membranes was found to be limited by the bulk ambipolar conductivity. At 1173 K and oxygen partial pressures 10 to 1.01×10⁵ Pa, ionic conductivity of intergrowth strontium ferrite increases with reducing oxygen pressure, indicating that the vacancy migration mechanism provides a greater contribution to ionic transport in comparison with the interstitial diffusion. The ion transference numbers of Sr₄Fe₆O_{13± δ} increase from 2.5×10^–4 to 1.9×10^–3 when the oxygen pressure decreases. Combined X-ray and neutron powder diffraction studies of oxygen-deficient Sr₄Fe₆O_{13– δ} showed an accumulation of oxygen vacancies in the non-perovskite layers, built of oxygen-iron polyhedra with pentacoordinated Fe cations. Both thermal and chemically-induced expansion of Sr₄Fe₆O_{13± δ} lattice have pronounced anisotropic character and are considerably lower than that typical for perovskite-type strontium ferrite. The average thermal expansion coefficients of Sr₄Fe₆O_{13± δ} ceramics at 770–1100 K in oxidizing and reducing atmospheres vary in the range (10.8–13.2)×10^–6 K^–1. Electronic Publication     

Protic conversion of nitrile into azavinylidene complexes of rhenium, a mechanistic theoretical study

The mechanism of the protonation of the rhenium nitrile chloro-complexes [ReCl(NCCH3)(PH3)4] (2), taken as models of the real systems [ReCl(NCR)(dppe)(2)] (dppe = Ph2PCH2CH2PPh2), leading to the azavinylidene products [ReCl(NC(H)CH3)(PH3)4]+ (3) was investigated by theoretical methods at the B3LYP level of theory. Electrostatic and molecular orbital arguments and thermodynamic, kinetic, and steric factors are analyzed and indicate that the chlorine atom is the most probable site of the initial proton attack, although the direct protonation of the nitrile carbon atom is also possible as a concurrent process. For the cis-isomer of 2, the initially formed chloro-protonated species cis-[Re(ClH)(NCCH3)(PH3)4]+ further converts to the azavinylidene cis-3 via either an acid-independent 1,4-proton shift or an acid-base catalyzed pathway involving a second protonation of the nitrile carbon atom to give cis-[Re(ClH)(NC(H)CH3)(PH3)4]2+ followed by elimination of the proton from the chlorine atom.