Acetylcholinesterase: mechanisms of covalent inhibition of wild-type and H447I mutant determined by computational analyses

The reaction mechanisms of two inhibitors TFK+ and TFK0 binding to both the wild-type and H447I mutant mouse acetylcholinesterase (mAChE) have been investigated by using a combined ab initio quantum mechanical/molecular mechanical (QM/MM) approach and classical molecular dynamics (MD) simulations. In the wild-type mAChE, the binding reactions of TFK+ and TFK0 are both spontaneous processes, which proceed through the nucleophilic addition of the Ser203-Ogamma to the carbonyl-C of TFK+ or TFK0, accompanied with a simultaneous proton transfer from Ser203 to His447. No barrier is found along the reaction paths, consistent with the experimental reaction rates approaching the diffusion-controlled limit. By contrast, TFK+ binding to the H447I mutant may proceed with a different reaction mechanism. A water molecule takes over the role of His447 and participates in the bond breaking and forming as a "charge relayer". Unlike in the wild-type mAChE case, Glu334, a conserved residue from the catalytic triad, acts as a catalytic base in the reaction. The calculated energy barrier for this reaction is about 8 kcal/mol. These predictions await experimental verification. In the case of the neutral ligand TFK0, however, multiple MD simulations on the TFK0/H447I complex reveal that none of the water molecules can be retained in the active site as a "catalytic" water. Furthermore, our alchemical free energy calculation also suggests that the binding of TFK0 to H447I is much weaker than that of TFK+. Taken together, our computational studies confirm that TFK0 is almost inactive in the H447I mutant and also provide detailed mechanistic insights into the experimental observations.     

Spectroscopic and electronic structure studies of intermediate X in ribonucleotide reductase R2 and two variants: a description of the FeIV-oxo bond in the FeIII-O-FeIV dimer

Spectroscopic and electronic structure studies of the class I Escherichia coli ribonucleotide reductase (RNR) intermediate X and three computationally derived model complexes are presented, compared, and evaluated to determine the electronic and geometric structure of the FeIII-FeIV active site of intermediate X. Rapid freeze-quench (RFQ) EPR, absorption, and MCD were used to trap intermediate X in R2 wild-type (WT) and two variants, W48A and Y122F/Y356F. RFQ-EPR spin quantitation was used to determine the relative contributions of intermediate X and radicals present, while RFQ-MCD was used to specifically probe the FeIII/FeIV active site, which displayed three FeIV d-d transitions between 16,700 and 22,600 cm(-1), two FeIV d-d spin-flip transitions between 23,500 and 24,300 cm(-1), and five oxo to FeIV and FeIII charge transfer (CT) transitions between 25,000 and 32,000 cm(-1). The FeIV d-d transitions were perturbed in the two variants, confirming that all three d-d transitions derive from the d-pi manifold. Furthermore, the FeIV d-pi splittings in the WT are too large to correlate with a bis-mu-oxo structure. The assignment of the FeIV d-d transitions in WT intermediate X best correlates with a bridged mu-oxo/mu-hydroxo [FeIII(mu-O)(mu-OH)FeIV] structure. The mu-oxo/mu-hydroxo core structure provides an important sigma/pi superexchange pathway, which is not present in the bis-mu-oxo structure, to promote facile electron transfer from Y122 to the remote FeIV through the bent oxo bridge, thereby generating the tyrosyl radical for catalysis.     

Determination of ascorbic acid in pharmaceutical dosage forms and urine by means of an oscillatory reaction system using the pulse perturbation technique

A simple and reliable method for the determination of ascorbic acid (AA) is proposed and validated. It is based on potentiometric monitoring of the concentration perturbations of an oscillatory reaction system in a stable nonequilibrium stationary state close to the bifurcation point. The response of the Bray–Liebhafsky (BL) oscillatory reaction as a matrix, to the perturbation by different concentrations of AA, is followed by a Pt electrode. The linear relationship between maximal potential shift and the logarithm of the amount of AA is obtained between 0.01 and 1.0 μmol. The sensitivity of the proposed method (as the limit of detection) is 0.009 μmol and the method has excellent sample throughput (30 samples per hour). The procedure was used for AA determination in pharmaceutical formulations and urine. The results are in agreement with those obtained using the official method. Some aspects of the possible mechanism of AA action on the BL oscillating chemical system are discussed.     

Calculus of functors,operad formality,and rational homology of embedding spaces

Let M be a smooth manifold and V a Euclidean space. Let [`(\textEmb)] \overline{{{\text{Emb}}}} (M,V) be the homotopy fiber of the map Emb(M,V) → Imm(M,V). This paper is about the rational homology of [`(\textEmb)] \overline{{{\text{Emb}}}} (M,V). We study it by applying embedding calculus and orthogonal calculus to the bifunctor (M,V)↦ HQ ∧ [`(\textEmb)] \overline{{{\text{Emb}}}} (M,V)₊. Our main theorem states that if

Homomorphism-Homogeneous Partially Ordered Sets

A structure is called homogeneous if every isomorphism between finite substructures of the structure extends to an automorphism of the structure. Recently, P. J. Cameron and J. Nešetřil introduced a relaxed version of homogeneity: we say that a structure is homomorphism-homogeneous if every homomorphism between finite substructures of the structure extends to an endomorphism of the structure. In this paper we characterize homomorphism-homogeneous partially ordered sets (where a homomorphism between partially ordered sets A and B is a mapping f : A →B satisfying ). We show that there are five types of homomorphism-homogeneous partially ordered sets: partially ordered sets whose connected components are chains; trees; dual trees; partially ordered sets which split into a tree and a dual tree; and X ₅-dense locally bounded partially ordered sets. Supported by the Ministry od Science and Environmental Protection of the Republic of Serbia, Grant No. 144017.     

The influence of daylight exposure on platelet 5-HT levels in patients with major depression and schizophrenia

Platelet serotonin (5-HT) can be used as a limited, peripheral model for the central 5-HT synaptosomes. Altered platelet 5-HT concentrations have been associated with psychiatric disorders like depression and schizophrenia. The aim of the present study was to compare platelet 5-HT concentrations during long, medium and short period of natural daylight exposure in a large number of medication-free male and female schizophrenic and depressed patients and sex-matched healthy controls. Platelet 5-HT concentration was determined spectrofluorimetrically in 240 (97 female, 143 male) schizophrenic and 258 (153 female, 105 male) nonpsychotic, nonsuicidal depressed medication-free patients and 328 (149 women, 179 men) healthy subjects during periods with short (<12), long (>12) and medium (average 12) hours of the natural daylight. Platelet 5-HT concentration was significantly lower in women compared to men in all groups. Healthy male subjects had significantly higher (p=0.011) platelet 5-HT concentrations during long compared to medium period. There were no significant differences (p>0.05) in platelet 5-HT concentration between different periods in healthy women. The significant increase in platelet 5-HT values were found in female (p=0.01) and male (p=0.029) depressed patients during long compared to short period. There were no significant associations between platelet 5-HT concentrations and different periods in both male and female schizophrenic patients. The results indicate the sex-related differences in the serotonergic system. The alterations of platelet 5-HT concentrations, observed across period with different durations of daylight exposure, point to a direct or indirect effect of light on peripheral 5-HT system that could be related to different sensitivity of the pineal gland to light and/or melatonin influence on 5-HT metabolism.     

Excessive excitation of hydrogen peroxide during oscillatory chemical evolution

The peculiar reaction dynamics of the Bray-Liebhafsky chemical oscillator is connected with the nonequilibrium periodic excitations of hydrogen peroxide embedded in a hydrogen-bonded water network. This was indicated by Raman spectroscopy that showed periodic, isothermal, and excessive excitation of the symmetric vibration of hydrogen peroxide. Since such an excessive excitation should be the result of a specific nonequilibrium energy distribution with the active participation of water, understanding this process can be of considerable importance in other systems in which water is the main constituent.     

Signature of the conformational preferences of small peptides: a theoretical investigation

An extensive computational study of the conformational preferences of N-acetylphenylalaninylamide (NAPA) is reported, including conformational and anharmonic frequency analyses, as well as calculations of excitation energies of the four NAPA conformers lowest in energy. Particular attention is paid to the influence of hydrogen-bonding interactions on the relative stability of the conformers, which was found to be very sensitive to both the level of quantum chemical computations and the anharmonic treatment of molecular vibrations. The assignments of the UV spectral peaks are well supported by the multireference CASSCF/MS-CASPT2 calculations. Upon consideration of the second-order M?ller-Plesset (MP2) and density functional theory (DFT) structures, overall energetics, and harmonic and anharmonic corrections, we found no conclusive theoretical evidence for the assumed conformational propensity of small model peptides toward extended beta-strand structures.     

Conversion of perhydropolysilazane into a SiOx network triggered by vacuum ultraviolet irradiation: access to flexible, transparent barrier coatings

The photochemical conversion of 200-500 nm layers of perhydropolysilazane --(SiH2-NH)n-- (PHPS) in the presence of oxygen into an SiOx network was studied. Different UV sources in the wavelength range of 160-240 nm, that is, 172 nm Xe2* and 222 nm KrCl* excimer, and 185 nm Hg low-pressure (HgLP) lamps were used for these purposes. The role of both ozone and O(1D) as well as of catalytic amounts of tertiary amines in the degradation process of PHPS and the formation of SiOx were studied. In this context, the kinetics of the entire reaction were elucidated and allowed both a continuous and discontinuous process to be established for the production of fully transparent, flexible barrier coatings. Barrier improvement factors (BIFs) of 400 were achieved with one single layer on 23 microm poly(ethyleneterephthalate) (PET), which translated into oxygen transmission rates (OTRs) of 0.20 cm3 m(-2) day(-1) bar(-1). Double layers prepared by this technique allowed the realization of OTRs of or=800.     

Possible existence of alkali metal orthocarbonates at high pressure

We investigate the possible existence of crystalline alkali metal orthocarbonates, A(4)CO(4), where A=Li, Na, K, Rb, and Cs. We study the equilibrium between the possible modifications of the orthocarbonate A(4)CO(4) and the binary mixture of the possible modifications of the alkali oxide A(2)O and those of the alkali metal carbonate A(2)CO(3) as function of pressure. In all cases, the orthocarbonate should be stable at sufficiently high pressure ranging from 22-32 GPa (Rb(4)CO(4)) to 200-220 GPa (Cs(4)CO(4)).