Molecular Wires with Controllable π-Delocalization Incorporating Redox-Triggered π-Conjugated Switching Units.

A perfluorobiphenyl-2,2'-diyl dication and its corresponding dihydrophenanthrene-type electron donor are interconvertible upon two-electron transfer. Redox-triggered C-C bond-formation/cleavage caused a drastic change in the torsion angle of the biphenyl unit. Thus, π-delocalization ON/OFF switching was observed as a change in the UV absorption upon electrolysis of the linearly extended analogue with two phenylethynyl groups.

Simultaneous computed tomographic angiography of the coronary and radial arteries.

Our aim was to assess a method of simultaneous computed tomographic (CT) angiography of the coronary and radial arteries without increasing contrast medium usage. Radial access is a standard approach in coronary interventions. However, radial artery puncture is difficult due to the small size of the radial artery and potential unexpected anomaly.

Assembly of an Axially Chiral Dynamic Redox System with a Perfluorobiphenyl Skeleton into Dumbbell- or Tripod-type Electron Donors.

The incorporation of F atoms endows a diethenylbiphenyl-based electron donor with configurational stability and SN Ar reactivity. The former enables the dynamic redox pair of (Rax)-1/(Rax ,R,R)-1(2+) to exhibit drastic UV/Vis and CD spectral changes upon electrolysis, whereas the latter makes it possible for (Rax)-1 to serve as a useful chiral synthon for the production of larger assemblies [(Rax ,Rax)-2 d,p,m and (Rax ,Rax ,Rax)-3] containing two or three dyrex units. These dyads and triad also exhibit a clean electrochiroptical response with isosbestic points owing to one-wave multi-electron transfer.

Induced preference for axial chirality in a triarylmethylium o,o-dimer upon complexation with natural γ-cyclodextrin: strong ECD signaling and fixation of supramolecular chirality to molecular chirality.

The ratio of the easily interconverting rotational isomers of biphenyl-2,2'-diylbis[bis(4-dimethylaminophenyl)methylium] (R)/(S)-1a(2+) can be biased to prefer an R configuration upon 1:1 complexation with γ-cyclodextrin in water. Through the reaction with Na(2)S, the preference of 1a(2+)@γ-CyD for an axial chirality of R can be fixed as the M-helicity of dihydrothiepin 2.

Diverse substrate recognition and hydrolysis mechanisms of human NUDT5.

Human NUDT5 (hNUDT5) hydrolyzes various modified nucleoside diphosphates including 8-oxo-dGDP, 8-oxo-dADP and ADP-ribose (ADPR). However, the structural basis of the broad substrate specificity remains unknown. Here, we report the crystal structures of hNUDT5 complexed with 8-oxo-dGDP and 8-oxo-dADP.

FT-IR spectroscopic studies on the molecular mechanism for substrate specificity/activation of medium-chain acyl-CoA dehydrogenase.

The interactions of acyl-CoA with medium-chain acyl-CoA dehydrogenases (MCADs) reconstituted with artificial FADs-i.e. 8-CN-, 7,8-Cl(2)-, 8-Cl-, 8-OCH(3)- and 8-NH(2)-FAD-were investigated by UV-visible absorption and FT-IR measurements.

Intramolecular and intermolecular perturbation on electronic state of FAD free in solution and bound to flavoproteins: FTIR spectroscopic study by using the C = O stretching vibrations as probes.

The intramolecular and intermolecular perturbation on the electronic state of FAD was investigated by FTIR spectroscopy by using the C=O stretching vibrations as probes in D(2)O solution. Natural and artificial FADs, i.e.

Theoretical study on charge-transfer interaction between acyl-CoA dehydrogenase and 3-thiaacyl-CoA using density functional method.

Acyl-CoA dehydrogenase forms a complex with a substrate analog, 3-thiaacyl-CoA, exhibiting a charge-transfer (CT) band. The structure of a complex model of oxidized lumiflavin with deprotonated 3-thiabutanoate ethylthioester designed for the above CT complex was fully optimized by means of density functional theory (DFT), the spatial arrangement being similar to the corresponding X-ray structure reported previously. The electrostatic interaction between flavin and an anionic ligand, therefore, plays a major role in determination of the arrangement of the CT complex.

Three-dimensional structure of rat-liver acyl-CoA oxidase in complex with a fatty acid: insights into substrate-recognition and reactivity toward molecular oxygen.

The three-dimensional structure of rat-liver acyl-CoA oxidase-II (ACO-II) in a complex with a C12-fatty acid was solved by the molecular replacement method based on the uncomplexed ACO-II structure. The crystalline form of the complex was obtained by cocrystallization of ACO-II with dodecanoyl-CoA. The crystalline complex possessed, in the active-site crevice, only the fatty acid moiety that had been formed through hydrolysis of the thioester bond.

Large-scale preparation of recombinant human calcitonin from a multimeric fusion protein produced in Escherichia coli.

In order to develop a large-scale, high-yield production process for human calcitonin (hCT) in Escherichia coli, a stable expression plasmid was constructed and the expressed protein was modified for efficient cleavage by protease. Multiple copies of a synthetic gene encoding hCT-Leu-Arg, a substrate for C-terminal amidation by carboxypeptidase Y (CPY), were inserted into the stable expression plasmid. Using this plasmid, the expression of a multimeric fusion protein was induced by shifting the temperature from 34 to 38 degrees C.

Hydrophobic core around tyrosine for human endothelin-1 investigated by photochemically induced dynamic nuclear polarization nuclear magnetic resonance and matrix-assisted laser desorption ionization time-of-flight mass spectrometry.

Human endothelin-1 (ET-1) is a potent cardiovascular bioactive peptide. Its activity is based on the C-terminal residues, e.g.

Protective immunity of SpaA-antigen producing Lactococcus lactis against Erysipelothrix rhusiopathiae infection.

Aims: To develop an economical, safe and simple vaccination system against swine erysipelas using SpaA-antigen producing Lactococcus lactis.

Methods And Results: The spaA gene of Erysipelothrix rhusiopathiae was inserted into a shuttle plasmid pSECE1 to construct pSECE1.3.

Distributed computing and NMR constraint-based high-resolution structure determination: applied for bioactive Peptide endothelin-1 to determine C-terminal folding.

Distributed computing has been implemented to the solution structure determination of endothelin-1 to evaluate efficiency of the method for NMR constraint-based structure calculations. A key target of the investigation was determination of the C-terminal folding of the peptide, which had been dispersed in previous studies of NMR, despite its pharmacological significances. With use of tens of thousands of random initial structures to explore the conformational space comprehensively, we determined high-resolution structures with good convergences of C-terminal as well as previously defined N-terminal structures.

Molecular mechanism of the drop in the pKa of a substrate analog bound to medium-chain acyl-CoA dehydrogenase: implications for substrate activation.

The pKa value of a substrate analogue 3-thiaoctanoyl-CoA at alphaC-H is known to drop from ca. 16 in the free state to 5-6 upon binding to medium-chain acyl-CoA dehydrogenase (MCAD). The molecular mechanism underlying this phenomenon was investigated by taking advantage of artificial FADs, i.

Structure of the transition state analog of medium-chain acyl-CoA dehydrogenase. Crystallographic and molecular orbital studies on the charge-transfer complex of medium-chain acyl-CoA dehydrogenase with 3-thiaoctanoyl-CoA.

The flavoenzyme medium-chain acyl-CoA dehydrogenase (MCAD) eliminates the alpha-proton of the substrate analog, 3-thiaoctanoyl-CoA (3S-C8-CoA), to form a charge-transfer complex with deprotonated 3S-C8-CoA. This complex can simulate the metastable reaction intermediate immediately after the alpha-proton elimination of a substrate and before the beta-hydrogen transfer as a hydride, and is therefore regarded as a transition-state analog. The crystalline complex was obtained by co-crystallizing MCAD in the oxidized form with 3S-C8-CoA.

Fourier transform IR study of aggregational behavior of N-acetyl-L- and N-butyloxycarbonyl-L-glutamic acid oligomeric benzyl esters in dioxane and benzene: beta-turn --> antiparallel beta-sheet transition.

Oligomeric N-acetyl-L-glutamic acid benzyl esters (AN(p)Z) with exact residue numbers (N(p) = 2, 3, 4, and 5) and N-butyloxycarbonyl-L-glutamic acid benzyl esters (BOCN(p)Z) with exact residue numbers (N(p) = 4, 5, 6, and 8) are synthesized by a stepwise procedure in a liquid phase. The aggregational behavior of these oligomeric molecules in dioxane and benzene is examined by Fourier transform IR spectra. In particular, the concentration dependence of the IR spectra for the AN(p)Z solutions with N(p) values of 4 (A4Z) and 5 (A5Z) shows that the predominant antiparallel beta-sheet structure is stabilized above the critical aggregation concentration (cac), while other conformations including beta-turns may coexist below the cac.

Three-dimensional structure of the flavoenzyme acyl-CoA oxidase-II from rat liver, the peroxisomal counterpart of mitochondrial acyl-CoA dehydrogenase.

Acyl-CoA oxidase (ACO) catalyzes the first and rate-determining step of the peroxisomal beta-oxidation of fatty acids. The crystal structure of ACO-II, which is one of two forms of rat liver ACO (ACO-I and ACO-II), has been solved and refined to an R-factor of 20.6% at 2.

Effects of hydrogen bonds in association with flavin and substrate in flavoenzyme d-amino acid oxidase. The catalytic and structural roles of Gly313 and Thr317.

According to the three-dimensional structure of a porcine kidney D-amino acid oxidase-substrate (D-leucine) complex model, the G313 backbone carbonyl recognizes the substrate amino group by hydrogen bonding and the side-chain hydroxyl of T317 forms a hydrogen bond with C(2)=O of the flavin moiety of FAD [Miura et al. (1997) J. Biochem.

Crystallization and preliminary X-ray characterization of rat liver acyl-CoA oxidase.

A recombinant form of the flavoenzyme acyl-CoA oxidase from rat liver has been crystallized by the hanging-drop vapour-diffusion technique using PEG 20 000 as a precipitating agent. The crystals grew as yellow prisms, with unit-cell parameters a = 71.05, b = 87.

Mechanism for the recognition and activation of substrate in medium-chain acyl-CoA dehydrogenase.

The mechanism underlying the recognition and activation of the substrate for medium-chain acyl-CoA dehydrogenase (MCAD) was spectroscopically investigated using 3-thiaacyl-CoAs as substrate analogs. The complex of MCAD with 3-thiaoctanoyl-CoA (3-thia-C8-CoA) exhibited a charge-transfer (CT) band with a molar extinction coefficient of epsilon808 = 9.1 mM-1.

Folding motifs induced and stabilized by distinct cystine frameworks.

Bioactive peptides of different sources and biological functionalities, like endothelins, sarafotoxins, bee and scorpion venom toxins, contain a consensus cystine framework, Cys-(X)1-Cys/Cys-(X)3-Cys, which has been found to induce and stabilize a homologous folding motif named the cystine-stabilized alpha-helix (CSH). This is composed of an alpha-helical segment spanning the Cys-(X)3-Cys sequence portion that is crosslinked by two disulfide bridges to the sequence portion Cys-(X)1-Cys, itself folded in an extended beta-strand type structure. Search for sequence homologies of peptides and proteins in the SWISS-PROT and PDB data banks provided additional multiple examples of this type of cystine framework in serine proteinase inhibitors, in insect and plant defense proteins, as well as in members of the growth factor family with the cystine-knot.

Spectroscopic studies of rat liver acyl-CoA oxidase with reference to recognition and activation of substrate.

Two forms of rat peroxisomal acyl-CoA oxidase (ACO-I and -II) interact with the substrate analogs, 3-ketoacyl-CoAs, forming a complex characterized by the so-called charge-transfer (CT) band around 575 nm in the absorption spectra. The CT band of ACO-I exhibited a broad dependency on the acyl chain-length from C4 to C16, whereas that of ACO-II showed increased intensity with a longer acyl chain to reach a maximum with a chain-length of C12. These chain-length dependencies of the CT bands were compared with those of the enzymatic activities reported previously [Setoyama et al.

Functional expression of two forms of rat acyl-CoA oxidase and their substrate specificities.

Using the reverse transcription of RNA followed by the polymerase chain reaction, we cloned the cDNAs for the rat acyl-CoA oxidases I and II, which are produced by alternative splicing from a single gene, and developed a system for their expression in Escherichia coli. The homogeneous preparations of these enzymes, without proteolytic procession, showed oxidase activity with acyl-CoAs having various acyl-chain lengths. The two types of the enzyme exhibited different substrate specificities with respect to the acyl-chain length, acyl-CoA oxidase I showing the optimum activity at shorter chain-length relative to acyl-CoA oxidase II.

[Lys(-2)-Arg(-1)]endothelin-1 solution structure by two-dimensional 1H-NMR: possible involvement of electrostatic interactions in native disulfide bridge formation and in biological activity decrease.

Addition of the Lys(-2)-Arg(-1) dipeptide, present in the precursor protein, to the N-terminus of endothelin-1 (ET-1), to form a 23-residue peptide (KR-ET-1) has been shown to greatly improve formation of native disulfide bridges and to dramatically decrease biological activity. Conformational analysis was carried out on this peptide. During protonation of the carboxyl groups, CD spectra showed a decrease in the helical contribution, and NMR spectra displayed strong chemical shift modifications, suggesting the importance of electrostatic interactions in the KR-ET-1 conformation.

Design, synthesis, and conformation of a model peptide of endothelin with cystine-stabilized alpha-helix motif.

A model 16-peptide of endothelin-1 (MET-1), which has the minimized sequence homology to the corresponding part of endothelin-1 (ET-1), was designed to confirm the cystine-stabilized alpha-helix motif. The model structure consists of an extended structure, a beta-turn part, and an alpha-helix structure that is stabilized by two disulfide bonds. The alpha-helix segment was designed to emphasize the amphiphilic nature.