Thermodynamical studies of designed ligands binding to Cel7A using partial-filling capillary electrophoresis.

A convenient experimental method for thermodynamical studies based on partial-filling affinity CE is presented. The advantages of this approach are the possibility to determine binding energies from relatively weak interactions as well as the small amounts of samples consumed. In order to explore the affinity and selectivity of the cellobiohydrolase Cel7A, a number of propranolol analogues were recently designed. The affinities of a selection of these ligands were determined in the temperature interval 15-40 degrees C, and DeltaG degrees , DeltaH degrees and DeltaS degrees were obtained by means of Van't Hoff plots. Through these experiments, the importance of the entropy contribution in the complexation between the ligands and Cel7A has been demonstrated.

pH dependency of ligand binding to cellobiohydrolase 1 (Cel7A). Affinity, selectivity and inhibition for designed propranolol analogues.

The affinity and enantioselectivity have been determined for designed propranolol derivatives as ligands for Cel7A by capillary electrophoresis (CE) at pH 7.0. These results have been compared to measurements at pH 5.0. In agreement with previous studies, the affinity increased at the higher pH. However, the affinity was not as dependent of the ligand structure at pH 7.0 as at pH 5.0, and the selectivity was generally decreased. Instead, at pH 7.0, the changes in binding were mainly dependent on the presence of additional dihydroxyl groups, indicating an increased importance of the electrostatic interactions. To evaluate the pH dependent variations in binding, changes in both the ligand and in the enzyme had to be taken into account. To ensure that the ligands had the same charge in all measurements, pKa-values of all compounds were determined. The ligand-protein interaction has also been studied by inhibition experiments at both pHs to evaluate the specific binding to the active site when competing with the substrate p-nitrophenyl lactoside (pNPL). With support of docking computations we propose a hypothesis on the effect of the ligand structure and pH dependency of the binding and selectivity of amino alcohols to Cel7A.

New propranolol analogues: binding and chiral discrimination by cellobiohydrolase Cel7A.

Novel propranolol analogues have been designed and synthesised and their enantioselective binding to the cellulose degrading enzyme, Cel7A, has been evaluated. Affinity and enantioselectivity have been determined by capillary electrophoresis experiments. Ligands with significantly improved affinity and selectivity have been obtained and an analysis of the results has led to insights concerning the relation between the changes in ligand structure and selectivity as well as affinity to the protein.

Spirobicyclo[2.2.2]octane derivatives: mimetics of baccatin III and paclitaxel (Taxol).

The formylated spirobyclic alcohol was computer modeled to be a mimetic of paclitaxel. In this model, the formyl group was used as a truncated paclitaxel side chain in order to reduce the computational work. Compound , carrying the paclitaxel side chain, was synthesized in six steps from optically active 1,3-diketone . Microtubule stabilization was not observed for , indicating that the model needs to be adjusted.

Design and synthesis of a C2-symmetric self-complementary hydrogen-bonding cleft molecule based on the bicyclo[3.3.1]nonane and 4-oxo-5-azaindole framework. formation of channels and inclusion complexes in the solid state.

The synthesis of a C2-symmetric cleft molecule 2 based on the fused framework between bicyclo[3.3.1]nonane and 4-oxo-5-azaindole, incorporating a self-complementary hydrogen-bonding motif, in both racemic and enantiomerically pure forms is reported. This cleft molecule is reminiscent of analogues of Tröger's base though with different cleft dimensions and tilt angles. The framework of 2 provides a building block for the construction of self-assembled hydrogen-bonded supramolecular structures. The solid-state structure of 2 is highly influenced by the limited solubility of (+/-)-2 and (-)-2. The solvents interact with the potential hydrogen-bonding motifs of (+/-)-2 and (-)-2, forming different three-dimensional structures as revealed by X-ray diffraction analysis. In the solid state (+/-)-(2)2 x 5DMF forms hydrogen-bonded pleated band structures that build up three-dimensional pens between adjacent bands in which two molecules of DMF are trapped. In contrast, the aggregate obtained from (-)-2, (-)-2 x 2AcOH, showed infinite bands of complex constitution.

Stereochemical variations on the colchicine motif. Peracid oxidation of thiocolchicone. Synthesis, conformation and inhibition of microtubule assembly.

When 7-oxodesacetamidothiocolchicine (1) was treated with various peroxides in order to afford a Baeyer-Villiger rearrangement, a complex mixture of products was formed, which included the sulfoxide, (2) and sulfone, (3). When peracetic acid was used two additional products were formed; a C-ring lactone (4) and a ring-contracted allocolchicine derivative (5). The sulfoxide (2) was semi-preparatively resolved into enantiomers by chromatography on microcrystalline triacetylcellulose. Rotational barriers around the A-C pivot bond of, and were determined by dynamic 1H NMR analysis. The compounds, and exhibit moderate inhibition of tubulin polymerization, according to in vitro turbidity studies, whereas was inactive.

Inter- and intramolecular potential for the N-formylglycinamide-water system. A comparison between theoretical modeling and empirical force fields.

An intramolecular NEMO potential is presented for the N-formylglycinamide molecule together with an intermolecular potential for the N-formylglycinamide-water system. The intramolecular N-formylglycinamide potential can be used as a building block for the backbone of polypeptides and proteins. Two intramolecular minima have been obtained. One, denoted as C5, is stabilized by a hydrogen bonded five member ring, and the other, denoted as C7, corresponds to a seven membered ring. The interaction between one water molecule and the N-formylglycinamide system is also studied and compared with Hartree-Fock SCF calculations and with the results obtained for some of the more commonly used force fields. The agreement between the NEMO and SCF energies for the complexes is in general superior to that of the other force fields. In the C7 region the surfaces obtained from the intramolecular part of the commonly used force fields are too flat compared to the NEMO potential and the ab initio calculations. We further analyze the possibility of using a charge distribution obtained from one conformation to describe the charge distribution of other conformations. We have found that the use of polarizabilities and generic dipoles can model most of the changes in charge density due to the different geometry of the new conformations, but that one can expect additional errors in the interaction energies that are of the order of 1 kcal/mol.

New approach to biomimetic transamination using bifunctional [1,3]-proton transfer catalysis in thioxanthenyl dioxide imines.

A pyridoxamine equivalent, 9-aminothioxanthene 10,10-dioxide, has been designed that is capable of affording transamination in good to excellent yields of natural as well as artificial amino acids. Amidines and guanidines in catalytic amounts were capable of performing [1,3]-proton transfer in the imines under mild conditions, whereas various simple amines failed. The use of chiral catalysts resulted in modest asymmetric induction (ee < or = 45%). The electronic dependence in para-substituted phenyl glyoxylate imines, isotope effects, and computational studies support a stepwise, bifunctional mechanism for amidine and guanidine catalysts. Attempts toward an autocatalytic model system are described.