Natural and engineered plasmin inhibitors : applications and design strategies

Plasmin is the primary enzyme responsible for dissolution of fibrin in the circulatory system. Plasminogen, the zymogen of plasmin is expressed ubiquitously in the human body [1], with the predominant source being the liver [2, 3]. Plasminogen is produced as an 810 amino acid protein with a 19 amino acid leader peptide, which is cleaved during secretion to produce the mature 791 amino acid one-chain zymogen. This is converted to plasmin by cleavage of the Arg561 - Val562 scissile bond [4], resulting in an active protease consisting of two disulfide linked chains. The amino-terminal heavy chain (residues Glu1-Arg561) is comprised of a plasminogen/apple/nematode (PAN) domain [5] and five kringle domains of approximately equal size [6] while the light chain (residues Val562-Asn791) contains a serine protease domain homologous to trypsin with a catalytic triad comprising His603, Asp646 and Ser741 [7]. Both plasmin and plasminogen occur in two forms, full length and a Lys77-Lys78 activated variant produced through self catalysis (Figure 1). The former exists in a tight conformation through binding of Lys50 and/or Lys62 to kringle domain 5 [8, 9] while Lys78-plasminogen assumes a more relaxed conformation rendering it more susceptible to plasmin conversion [10, 11].

Supported gold nanoparticles as photocatalysts utilising the full solar spectrum for organic synthesis

This thesis is an innovative study for organic synthesis using supported gold nanoparticles as photocatalysts under visible light irradiation. It especially examines a novel green process for efficient hydroamination of alkynes with amines. The investigation of other traditional reduction and oxidation reactions also adds significantly to the knowledge of gold nanoparticles and titania nanofibres as photocatalysts for organic synthesis.

Technological Advances in Geotextiles

The role of polymer chemistry (pure and applied sciences) is very prominent in the world of science today, but it is heading away from polymers and polymer blends towards composites and nanocomposites. This allows for the creation of new materials with unique properties and new possibilities which is the subject of this new book.

Rapid match-searching for gene silencing assessment

Motivation: Gene silencing, also called RNA interference, requires reliable assessment of silencer impacts. A critical task is to find matches between silencer oligomers and sites in the genome, in accordance with one-to-many matching rules (G-U matching, with provision for mismatches). Fast search algorithms are required to support silencer impact assessments in procedures for designing effective silencer sequences.Results: The article presents a matching algorithm and data structures specialized for matching searches, including a kernel procedure that addresses a Boolean version of the database task called the skyline search. Besides exact matches, the algorithm is extended to allow for the location-specific mismatches applicable in plants. Computational tests show that the algorithm is significantly faster than suffix-tree alternatives. © The Author 2010. Published by Oxford University Press. All rights reserved.

Direct iodination of isoindolines and isoindoline nitroxides as precursors to functionalized nitroxides

A new method for the direct aryl iodination of isoindolines and isoindoline nitroxides which utilizes periodic acid and potassium iodide in sulfuric acid is presented. Di-iodo functionalized tetramethyl and tetraethyl isoindolines and a di-iodo tetramethyl isoindoline nitroxide were prepared in high yield (70-82%). The analogous mono-iodo species were afforded in modest yield (34-48%). Iodinated nitrones were also obtained from a tetraethyl isoindoline nitroxide.

Structures of μOconotoxins from Conusmarmoreus. Inhibitors of tetrodotoxin (TTX)-sensitive and TTX-resistant sodium channels in mammalian sensory neurons

The μO-conotoxins are an intriguing class of conotoxins targeting various voltage-dependent sodium channels and molluscan calcium channels. In the current study, we have shown MrVIA and MrVIB to be the first known peptidic inhibitors of the transient tetrodotoxin-resistant (TTX-R) Na+ current in rat dorsal root ganglion neurons, in addition to inhibiting tetrodotoxin-sensitive Na+ currents. Human TTX-R sodium channels are a therapeutic target for indications such as pain, highlighting the importance of the μO-conotoxins as potential leads for drug development. Furthermore, we have used NMR spectroscopy to provide the first structural information on this class of conotoxins. MrVIA and MrVIB are hydrophobic peptides that aggregate in aqueous solution but were solubilized in 50% acetonitrile/water. The three-dimensional structure of MrVIB consists of a small β-sheet and a cystine knot arrangement of the three-disulfide bonds. It contains four backbone “loops” between successive cysteine residues that are exposed to the solvent to varying degrees. The largest of these, loop 2, is the most disordered part of the molecule, most likely due to flexibility in solution. This disorder is the most striking difference between the structures of MrVIB and the known δ- and ω-conotoxins, which along with the μO-conotoxins are members of the O superfamily. Loop 2 of ω-conotoxins has previously been shown to contain residues critical for binding to voltage-gated calcium channels, and it is interesting to speculate that the flexibility observed in MrVIB may accommodate binding to both sodium and molluscan calcium channels.

Diporphyrins linked by two-atom bridges synthestic, structural and theoretical studies

The syntheses, properties and electronic structures of a series of porphyrin dimers connected by two-atom bridges were compared. The study found that an azo linker results in the most efficient electronic communication between the two porphyrin rings, and is the superior connector for dimers, trimers and oligomers in the design of nonlinear optical materials. This has implications for the design of molecular probes and sensors, photodynamic therapy, microfabrication, and three-dimensional optical data storage. The research led to the synthesis of a number of new porphyrin monomers and dimers, which were characterised using structural, spectroscopic and spectrometric techniques.

Synthesis of novel inhibitors blocking Wnt signaling downstream of β-Catenin

Large scale screening of libraries consisting of natural and small molecules led to the identification of many small molecule inhibitors repressing Wnt/β-Catenin signaling. However, targeted synthesis of novel Wnt pathway inhibitors has been rarely described. We developed a modular and expedient way to create the aromatic ring system with an aliphatic ring in between. Our synthesis opens up the possibility, in principle, to substitute all positions at the ring system with any desired substituent. Here, we tested five different haloquinone analogs carrying methoxy- and hydroxy-groups at different positions. Bona fide Wnt activity assays in cell culture and in Xenopus embryos revealed that two of these compounds act as potent inhibitors of aberrant activated Wnt/β-Catenin signaling.

Gas-phase reactions of aryl radicals with 2-butyne : an experimental and theoretical investigation employing the N-methyl-pyridinium-4-yl radical cation

Aromatic radicals form in a variety of reacting gas-phase systems, where their molecular weight growth reactions with unsaturated hydrocarbons are of considerable importance. We have investigated the ion-molecule reaction of the aromatic distonic N-methyl-pyridinium-4-yl (NMP) radical cation with 2-butyne (CH3C CCH3) using ion trap mass spectrometry. Comparison is made to high-level ab initio energy surfaces for the reaction of NMP and for the neutral phenyl radical system. The NMP radical cation reacts rapidly with 2-butyne at ambient temperature, due to the apparent absence of any barrier. The activated vinyl radical adduct predominantly dissociates via loss of a H atom, with lesser amounts of CH3 loss. High-resolution Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometry allows us to identify small quantities of the collisionally deactivated reaction adduct. Statistical reaction rate theory calculations (master equation/RRKM theory) on the NMP + 2-butyne system support our experimental findings, and indicate a mechanism that predominantly involves an allylic resonance-stabilized radical formed via H atom shuttling between the aromatic ring and the C-4 side-chain, followed by cyclization and/or low-energy H atom beta-scission reactions. A similar mechanism is demonstrated for the neutral phenyl radical (Ph center dot)+2-butyne reaction, forming products that include 3-methylindene. The collisionally deactivated reaction adduct is predicted to be quenched in the form of a resonance-stabilized methylphenylallyl radical. Experiments using a 2,5-dichloro substituted methyl-pyridiniumyl radical cation revealed that in this case CH3 loss from the 2-butyne adduct is favoured over H atom loss, verifying the key role of ortho H atoms, and the shuttling mechanism, in the reactions of aromatic radicals with alkynes. As well as being useful phenyl radical analogues, pyridiniumyl radical cations may form in the ionosphere of Titan, where they could undergo rapid molecular weight growth reactions to yield polycyclic aromatic nitrogen hydrocarbons (PANHs).

Identification, functional expression and kinetic analysis of two primary amine oxidases from Rhodococcus opacus

Two native copper-containing amine oxidases (EC 1.4.3.21) have been isolated from Rhodococcus opacus and reveal phenotypic plasticity and catalytic activity with respect to structurally diverse natural and synthetic amines. Altering the amine growth substrate has enabled tailored and targeted oxidase upreg-ulation, which with subsequent treatment by precipitation, ion exchange and gel filtration, achieved a 90–150 fold purification. MALDI-TOF mass spectrometric and genomic analysis has indicated multiple gene activation with complex biodegradation pathways and regulatory mechanisms. Additional post-purification characterisation has drawn on the use of carbonyl reagent and chelating agent inhibitors. Michaelis–Menten kinetics for common aliphatic and aromatic amine substrates and several structural analogues demonstrated a broad specificity and high affinity with Michaelis constants (K M) ranging from 0.1 to 0.9 mM for C 1 –C 5 aliphatic mono-amines and <0.2 mM for a range of aromatic amines. Potential exploitation of the enzymatic versatility of the two isolated oxidases in biosensing and bioprocessing is discussed.

Construction of interstellar cumulenes and heterocumulenes : Mass spectrometric studies

The last few years have brought an increasing interest in the chemistry of rite interstellar and circumstellar environs. Many of the molecular species discovered in remote galactic regions have been dubbed 'non-terrestrial' because of their unique structures (Thaddeus et al, 1993). These findings have provided a challenge to chemists in many differing fields to attempt to generate these unusual species in the laboratory of particular recent interest have been the unsaturated hydrocarbon families, CnH and CnH2, which have been pursued by a number of diverse methodologies. A wine range of heterocumulenes, including CnO, HCnO, CnN, HCnN, CnS, HCnS, CnSi and HCnSi have also provided intriguing targets for laboratory experiments. Strictly the term cumulene refers to a class of compounds that possess a series of adjacent double bonds, with allene representing the simplest example (H2C=C=CH2). However for many of the non-terrestrial molecules presented here, the carbon chain cannot be described in terms of a single simple valence structure, and so we use the terms cumulene and heterocumulene in a more general sense: to describe molecular species that contain an unsaturated polycarbon chain. Mass spectrometry has proved an invaluable tool in the quest for interstellar cumulenes and heterocumulenes in the laboratory it has the ability in its many forms, to (i) generate charged analogs of these species in the gas phase, (ii) probe their connectivity, ion chemistry, and thermochemistry, and (iii) in some cases, elucidate the neutrals themselves. Here, we will discuss the progress of these studies to this time. (C) 1999 John Wiley & Sons, Inc.

Mechanism‐based selection of a potent kallikrein‐related peptidase 7 inhibitor from a versatile library based on the sunflower trypsin inhibitor SFTI‐1

Potent and specific enzyme inhibition is a key goal in the development of therapeutic inhibitors targeting proteolytic activity. The backbone-cyclized peptide, Sunflower Trypsin Inhibitor (SFTI-1) affords a scaffold that can be engineered to achieve both these aims. SFTI-1's mechanism of inhibition is unusual in that it shows fast-on/slow-off kinetics driven by cleavage and religation of a scissile bond. This phenomenon was used to select a nanomolar inhibitor of kallikrein-related peptidase 7 (KLK7) from a versatile library of SFTI variants with diversity tailored to exploit distinctive surfaces present in the active site of serine proteases. Inhibitor selection was achieved through the use of size exclusion chromatography to separate protease/inhibitor complexes from unbound inhibitors followed by inhibitor identification according to molecular mass ascertained by mass spectrometry. This approach identified a single dominant inhibitor species with molecular weight of 1562.4 Da, which is consistent with the SFTI variant SFTI-WCTF. Once synthesized individually this inhibitor showed an IC50 of 173.9 ± 7.6 nM against chromogenic substrates and could block protein proteolysis. Molecular modeling analysis suggested that selection of SFTI-WCTF was driven by specific aromatic interactions and stabilized by an enhanced internal hydrogen bonding network. This approach provides a robust and rapid route to inhibitor selection and design.

4-Methoxyanilinium 2-carboxy-4,5-dichlorobenzoate

In the structure of the title salt C7H10NO+ C8H3Cl2O4- the benzene planes of the cation and anion are essentially parallel [inter-ring dihedral angle 4.8(2)deg]. In the anion the carboxylic acid and carboxylate groups make dihedral angles of 19.0(2) and 79.5(2)\%, respectively, with the benzene ring. Aminium N-H...O, carboxylic acid O-H...O and weak aromatic C-H...O hydrogen-bonding associations with carboxyl O-atom acceptors together with cation-anion pi-pi ring interactions [minimum ring centroid separation = 3.734(3)Ang] give a two-dimensional sheet structure which lies parallel to (001).

4-(4-Aminophenylsulfonyl)anilinium toluene-4-sulfonate

In the title p-toluenesulfonate salt of the drug dapsone, C12H13N2O2S+ C7H7O3S-, the dihedral angle between the two aromatic rings of the dapsone monocation is 70.19(17)deg. and those between these rings and that of the p-toluenesulfonate anion are 72.34(17) and 46.43(17)deg. All amine and aminium H-atoms are involved in intermolecular N-H...O hydrogen-bonding associations with sulfonyl O-atom acceptors as well as one of the sulfone O-atoms, giving a three-dimensional structure.

Proton-transfer and non-transfer compounds of the multi-purpose drug dapsone [4-(4-Aminophenylsulfonyl)aniline] with 3,5-dinitrosalicylic acid and 5-nitroisophthalic acid

The structures of the compounds from the reaction of the drug dapsone [4-(4-aminophenylsulfonyl)aniline] with 3,5-dinitrosalicylic acid, the salt hydrate [4-(4-aminohenylsulfonyl)anilinium 2-carboxy-4,6-dinitrophenolate monohydrate] (1) and the 1:1 adduct with 5-nitroisophthalic acid [4-(4-aminophenylsulfonyl)aniline 5-nitrobenzene-1,3-dicarboxylic acid] (2) have been determined. Crystals of 1 are triclinic, space group P-1, with unit cell dimensions a = 8.2043(3), b = 11.4000(6), c = 11.8261(6)Å, α = 110.891(5), β = 91.927(3), γ = 98.590(4)deg. and Z = 4. Compound 2 is orthorhombic, space group Pbcn, with unit cell dimensions a = 20.2662(6), b = 12.7161(4), c = 15.9423(5)Å and Z = 8. In 1, intermolecular analinium N-H…O and water O-H…O and O-H…N hydrogen-bonding interactions with sulfone, carboxyl, phenolate and nitro O-atom and aniline N-atom acceptors give a two-dimensional layered structure. With 2, the intermolecular interactions involve both aniline N-H…O and carboxylic acid O-H…O and O-H…N hydrogen bonds to sulfone, carboxyl, nitro and aniline acceptors, giving a three-dimensional network structure. In both structures π--π aromatic ring associations are present.