Binding energies of tyrosine kinase inhibitors: Error assessment of computational methods for imatinib and nilotinib binding

The binding energies of imatinib and nilotinib to tyrosine kinase have been determined by quantum mechanical (QM) computations, and compared with literature binding energy studies using molecular mechanics (MM). The potential errors in the computational methods include these critical factors:

• •Errors in X-ray structures such as structural distortions and steric clashes give unrealistically high van der Waals energies, and erroneous binding energies.
• •MM optimization gives a very different configuration to the QM optimization for nilotinib, whereas the imatinib ion gives similar configurations
• •Solvation energies are a major component of the overall binding energy. The QM based solvent model (PCM/SMD) gives different values from those used in the implicit PBSA solvent MM models. A major error in inhibitor—kinase binding lies in the non-polar solvation terms.
• •Solvent transfer free energies and the required empirical solvent accessible surface area factors for nilotinib and imatinib ion to give the transfer free energies have been reverse calculated. These values differ from those used in the MM PBSA studies.
• •An intertwined desolvation—conformational binding selectivity process is a balance of thermodynamic desolvation and intramolecular conformational kinetic control.
• •The configurational entropies (TΔS) are minor error sources.

     

Effects of static magnetic fields of erythrocyte rheology

The orientation of normal erythrocytes in a uniform static magnetic field (8 T maximum) has been investigated microscopically and photometrically.

• 1.(1) The intact erythrocytes were oriented with their disk planes parallel to the magnetic field because of the diamagnetism of the cell membrane components, particularly the transmembrane proteins (e.g., Band III, glycopholin) and the lipid bilayer.
• 2.(2) In contrast, the glutaraldehyde-fixed erythrocytes were oriented perpendicular to the field, perhaps because of the paramagnetism of the membrane-bound methemoglobin.
• 3.(3) The orientation was established within 5 s in a dilute suspension (5 × 10³cells μl⁻¹) as estimated from the change in light scattering after exposure to the magnetic field.

     

Physical Properties of Highly Active Liquor Containing Molybdate Solids

The reprocessing of irradiated nuclear fuel at Sellafield produces a nitric acid based Highly Active Liquor (HAL) waste. The liquor, containing fission products and process additives, is concentrated in an evaporator in order to reduce the volume and is then stored in Highly Active Storage Tanks (HASTs) prior to vitrification. Caesium phosphomolybdate (CPM) is precipitated during the evaporation process and can convert to zirconium molybdate (ZM) during storage.During Post Operational Clean Out (POCO) of the HASTs, it is expected that their highly active content will be reduced by repeated cycles of washing using nitric acid and other reagents. Initial washings are likely to have a chemical composition comparable to concentrated HAL, becoming more dilute during the wash-out process. It is expected that the wash-out process will also recover significant quantities of molybdate solids (ZM, CPM or a mixture) from the HASTs.In order to determine the processing challenges from such washings during POCO, the physical properties of varying concentrations of non-active HAL simulants containing molybdate solids have recently been measured by the UK's National Nuclear Laboratory. The following measurements are presented and discussed:

• •Particle size distribution
• •Density
• •Settling behaviour of solids
• •Voidage of settled sediment beds
• •Viscosity
• •Yield stress
• •Influence of ZM morphology on physical properties

     

899 — A potentiometric study on the redox properties of hemoglobin from Camelus dromedarius

A potentiometric study of the redox properties of hemoglobin from Camelus dromedarius has been performed, with the aim of having a better insight in the structure-function relationships of this protein; data on human hemoglobin have been included for comparative purposes. Analysis of the experimental data has shown:

• 1.the existence of intermediate conformers (neither R nor T) in the oxidized form of dromedary hemoglobin;
• 2.the amino acid groups involved in the oxidation Bohr effect of dromedary and human hemoglobins are likely to be the same; thus, only invariant amino acid residues seem to be involved in this effect.

     

Fourier transform infrared spectroscopic studies of the secondary structure and thermal denaturation of CaATPase from rabbit skeletal muscle

Fourier transform i.r. spectroscopy has been used to monitor structural alterations induced by thermal denaturation of the intrinsic membrane protein CaATPase in aqueous media. The protein has been isolated, purified and studied in five forms:

• 1.(i) In its native lipid environment after isolation from rabbit sarcoplasmic reticulum, both in H₂O and D₂O suspensions.
• 2.(ii) After both mild and extensive tryptic digestion has cleaved those residues external to the membrane bilayer.
• 3.(iii) Reconstituted in vesicle form with bovine brain sphingomyelin.

Fourier deconvolution techniques have been used to enhance the resolution of the intrinsically overlapped Amide I and Amide II spectral regions. Large spectral alterations apparent in the deconvoluted spectra occur in these regions upon thermal denaturation of the protein which are consistent with the formation of a large proportion of β-antiparallel sheet form. The alteration parallels the loss in ATPase activity. A mild tryptic digestion increases slightly the proportion of α-helix and/or random coil secondary structure. A thermal transition to a form containing a high proportion of β structure is still evident. Extensive tryptic digestion nearly abolishes the alpha helical plus random coil secondary structure, while producing a high proportion of β form which is resistant to further thermally induced structural alterations.Studies of CaATPase reconstituted into vesicles with bovine brain sphingomyelin reveal a higher proportion of β structure than the native enzyme, with further introduction of β structure on thermal denaturation. Both the utility of deconvolution techniques and the necessity for caution in their application are apparent from the current experiments.     

The adsorption of RNAse A,BSA and cytochrome c at the graphite powder|liquid interface using in parallel the adsorption isotherm plot and linear sweep voltammetry on graphite paste electrode

The adsorption of RNAse A, BSA and cytochrome c on graphite powder has been investigated using in parallel the adsorption isotherm plot and linear sweep voltammetry on a graphite paste electrode (g.p.e.). The principle of the latter depends on the determination by electrochemical oxidation of tyrosyl or tryptophan residues having access to the interface. Adsorption isotherms exhibit bimodal adsorption features with a cooperative phenomenon leading to the formation of a close-packed two dimensional surface phase. Protein molecules are adsorbed in a monolayer when the pH is different from the isoelectric point. This layer is built up by the entanglement of a double network:

• •A network of molecules irreversibly adsorbed side-on, where hydrophobic residues move from the core to the surface. The tertiary structure is likely altered. The new dimensions of these adsorbed molecules have been determined (surface area per molecule, thickness).
• •A network of molecules adsorbed end-on, lying between the irreversibly adsorbed molecules.

     

N–H stretching frequencies and the conformation of substituted ureas: an ab initio MO study

The dependence of N–H stretching-mode frequencies in representative di- and trialkyl ureas on the conformational state of the ureido group has been studied by ab initio MO calculations using HF/3-21G and HF/6-31G** basis sets. The molecules studied were 1,3-dimethylurea, 1-methyl-3,3-dimethylurea and 1-methyl-3,3-di-iso-propylurea. The principal conclusions from the ab initio results are:

• 1.the trans–trans conformer (N–H bonds trans to the CO bond) has N–H stretching bands with about 20–30 cm⁻¹ higher frequency than the respective cis–cis structure, in accord with earlier literature assignments based on experimental data;
• 2.the N–H stretching frequency interval in tri-substituted ureas is 15–20 cm⁻¹ higher than the N–H band position in the 1,3-disubstituted molecule studied, the effect being determined mostly by the higher N–H stretching force constant;
• 3.in the absence of the steric hindrance the stable rotameric forms of the ureido grouping are almost planar at HF/3-21G level of calculations, while HF/6-31G** calculations predict a slightly pyramidal structure at the nitrogen atoms in the trans–trans conformer;
• 4.in 1-methyl-3,3-di-iso-propylurea the steric influence of the two bulky iso-propyl groups cause a deviation from planarity of the N–H bond. The non-planar conformation is accompanied by a shift of the N–H stretching mode frequency towards higher values; and
• 5.the variations of the theoretically estimated N–H stretching-mode frequencies appear to be principally determined by changes in the N–H stretching force constants in the different molecules.

© 1997 Elsevier Science B.V.     

Latest Developments in Proton Conductors

Solid proton conductors are receiving considerable attention, stimulated by the need of pollution control and with the objective to develop future microprotonics. The latest striking developments in the study of proton conductors are presented:

• i)Hydrogen containing perovskites, with a discussion of the nature and location of hydrogen inprotonic and electronic high-temperature superconductors;
• ii)Novel forms of hydrogen in disordered and mixed conductors, with the problems of the vibrational proton transfer assistance assumed by some authors and the independent proton dynamics evidenced by inelastic neutron scattering;
• iii)Frequency dependent conductivity and dielectric relaxation, including thermally stimulated current experiments in hydrated synthetic and biological gels (seeds);
• iv)Proton conducting membranes as candidates to replace Nafion®-like materials in H₂ (methanol) fuel cells.

     

Synthese et caracterisation de fluorotellurates IV alcalins et d'ammonium partie I. Etude du pentafluorotellurate IV de lithium LiTeF5

Lithium pentafluorotellurate (IV) LiTeF₅ has been prepared by different methods:

• •slow evaporation of a 1/1 mixture of lithium fluoride and tellurium dioxide or tellurium tetrachloride in anhydrous hydrogen fluoride,
• •heating a 1/1 mixture of lithium fluoride and tellurium tetrafluoride. LiTeF₅ is orthorhombic

(a = 9,52 Å b = 9,12 Å c = 11, 60 Å)     

Recent studies of the interaction of rabbit dimeric IgA with its polymeric immunoglobulin receptor

Rabbit secretory components (SC) constitute a highly heteregeneous population of glycoprotein molecules that are present in secretions as free or bound forms to polymeric immunoglobulins (Ig). Two SC families are known, one of high molecular weight (⋍ 80 Kd) composed of five (perhaps six) domains related to Ig variable domains, and one of low molecular weight (⋍ 55 Kd) An account of our most recent experimental data is reviewed in this article. We have shown:

• 1.1) that both the high and low Mr SC families possess the same relative avidity for binding to dimeric IgA of the g-sublanguage;
• 2.2) that the first NH₂-terminal domain of SC derived from the high and low Mr polypeptides is necessary and sufficient for efficient non-covalent binding to dimeric IgA of the g-sublanguage;
• 3.3) that the low Mr SC polypeptide derives from the high Mr SC by the internal deletion of the entire second and third domains, suggesting that these domains are not involved in the binding reaction with polymeric Ig;
• 4.4) that the heterogeneity of rabbit secretory components is, in large part, due to the expression of several polymorphic forms (allotypes) susceptible to be recognized by specific alloantisera; the biochemical characterization of the three known SC allotypes (t61, t62 and t63) reveals that t62 and t63 are structurally very similar to each other and markedly divergent from the t61 homologue;
• 5.5) that by using non-cross-reactive alloantisera, the major immunodominant allotopes are confined within the COOH-terminal domains 3, 4 and 5 of SC;
• 6.6) that the location of the residues involved in the attachment of the carbohydrate unit within domain 1 varies according to the allotype: t61 is N-linked glycosylated at position 70, whereas about 75 % of t62 molecules are devoid of sugars; the remaining 25 % of t62 molecules are glycosylated at residue position 90; these oligosaccharide chain units are linked to asparagine residues in the acceptor site consensus sequence, Asn-X-Thr/Ser;
• 7.7) that the presence of the carbohydrate unit in domain 1 is not required for efficient binding of this domain to polymeric Ig: indeed, after enzymatic deglycosylation, domain 1 exhibits a relative binding avidity which is indistinguishable from that of the native glycosylated domain 1.

     

Application of radiation technology in biomedical materials; Fundamentals and applied

• 1.1) IMMOBILIZATION OF HEMOGLOBIN Hemoglobin has been immobilized into Poly Hema Matrix. To increase, Mechanical resistance, at first, CO was coodinated, after immobilization CO was eliminated by photo illumination by visible light from a W lamp and then O₂ was introduced. Oxygencoordiation ability was not damaged by immobilization.
• 2.2) REDUCTION MECHANISM OF ENZYME BY THE USE OF PULSE RADIOLYSIS Elementary process of Reduction Mechanism of Myoglobin, Hemoglobin, HRP and Cytochrome Oxidase were investigated in the time range of μsec≈nsec. In the of Cytochrome Oxidase, these are 4 metal ions inside of the Enzyme. The exact step of reduction of this enzyme was elucidated

.     

Calorimetric studies of CeCu AND CeMgCu alloys

Using different calorimeters, the following measurements have been carried out.

• 1.(i) The integral enthalpies of mixing of CeCu binary and some CeMgCu ternary liquid alloys,
• 2.(ii) The enthalpy of formation and the enthalpy of melting of the intermetallic compound CeCu₂.
• 3.(iii) The heat content of solid and liquid CeCu₂.

The observed concentration and temperature dependence of the measured values of the enthalpies of mixing as well as the glass-forming abilities of these alloys are explained on the basis of an association model.     

New experimental set-ups for studying nanoconfined water on the AILES beamline at SOLEIL

Three ensembles designed to investigate condensed matter in complex environments have been developed recently on the AILES beamline at SOLEIL. They have been exploited for studies aiming at understanding the properties of water molecules and their network in various confining systems, namely:

• -a hydration and temperature-controlled cell for the study of water confined in nanoporous Vycor,
• -a high pressure set-up allowing the study of the evolution of water molecules network trapped in Faujasite through the pressure-induced amorphisation of the matrix material,
• -a temperature resolved electrochemical cell used to record FIR difference spectra of metalloproteins interacting with water molecules.

By combining the high infrared flux and collimation of the AILES beamline with these optimized sample environments, it is possible to measure the infrared and THz spectra for minute quantities of samples in precise physical conditions.     

Über die Bestimmung der Richtung von Übergangsmomenten in länglichen Molekülen aus Messungen des IR-Dichroismus, 2. Mitt.:

Starting from a hypothesis developed byKratky about the motion of rod-like particles when the surrounding medium is extended, two methods for orientation of molecules will be treated:

1. Embedding of the molecules in plastic foils and stretching the foils.
2. For macromolecules: Spreading of jellies of the molecules to the foils and stretching the foils.

Relations between extension ratio and degree of orientation could be derived allowing to determine exactly the direction of transition moments.     

Hippurathydrolase,ein neues Enzym aus Mikroorganismen, 2. Mitt.: Enzymeigenschaften

Characteristic properties of the new enzymehippurate hydrolase are reported:

1. The Michaelis constant for hippurate hydrolysis is 5.5·10^?4 mole/l at 5° and pH 7.5. The specific activity of the purified enzyme preparation is 7.5 μmole/minute · mg enzyme protein at 37° and pH 7.5. The pH optimum is 7.5.
2. The effects of temperature on the reaction rate and on the stability of the enzyme were determined.
3. The effects of various inhibitors (HgCl₂, pCMB, EDTA, 8-hydroxyquinoline) and cations were studied. Participation of metal ions in the catalytic function ofhippurate hydrolase was not detectable.

     

Decay of (Fe1−xNix)0.96S DSC investigation

The decay of a monosulphide solid solution (mss) with the composition (Fe_1?1Ni_x)_0.96S was investigated by means of differential scanning calorimetry in the temperature range, from 20 to 305?C. Thermal effects of various natures were detected:

1. Ordering-disordering in the Fe-Ni sublattice near 100?C.
2. Pentlandite exsolution (exothermal peak); the peak temperature varies from 180 to 240?C and depends on the initial composition; the higher the Ni content, the lower the exsolution temperature.
3. Magnetic-paramagnetic transition. The transition temperature decreases down to 220?C as the Fe∶Ni ratio is decreased from 10∶0 to 4∶6.

Ni atoms are the defects in the magnetic ordering of themss generated by the Fe atoms in the metal sublattice. Thus, the driving force for pentlandite exsolution is the removal of Ni atoms from the magneticmss into the nonmagnetic pentlandite. This is the reason why the Fe∶Ni ratio in the generated pentlandite is much higher than that in the initialmss.     

Synthese et caracterisation de fluorotellurates IV alcalins et d'ammonium partie II: Etude des fluorotellurates d'ammonium NH4TeF5 et (NH4)2TeF6

New methods of synthesis:

• •reaction of aqueous or anhydrous HF with TeO₂ and NH₄F or NH₄HF₂, with (NH₄)₂TeCl₆
• •reaction of TeF₄ with NH₄F or NH₄HF₂
• •reaction of NH₄HF₂ with TeO₂

have been investigated for NH₄TeF₅ and a new compound (NH₄)₂TeF₆. The resulting compounds have been analysed and characterized.     

863 — Absorption and conversion of electric field energy by membrane bound atpases

Electric fields are ubiquitous in living organisms, yet, little is known of their effects on the function of enzymes and proteins. Recent experiments show that applied electric fields can induce active transport of K⁺, Rb⁺ and Ca²⁺ by transport-ATPases, as well as ATP synthesis by ATP-synthetases. In all cases, tightly sealed membrane vesicles have been essential.Here we present a kinetic model for the transduction of electric field energy based on three considerations:

• 1.The equilibrium between any two conformational states of a protein is susceptible to perturbation by an electric field so long as there exists a difference between the molar electric moments.
• 2.An integral membrane protein experiences an amplified electric field.
• 3.The transmembrane potential is maintained only with non-leaky vesicles.

This model consists of four steps to form a complete cycle (i.e. one enzyme turnover), namely, the conformational energy coupling (energy absorption), the substrate binding, the energy transduction (energy conversion), and the product release steps. The energy transduction step is mediated by reversal of the local field.Computer simulations show that this four state model converts electric field energy into chemical potential energy if the frequency and the strength of the applied field properly match the characteristics of the system. The model finds analogy in an electric rectifier, which conducts current only in certain directions. The results of simulation agree with the main features of the experimental data for voltage-stimulated active transport of Rb⁺via (Na,K)ATPase of erythrocytes.The concept is extended to the interpretation of data for the voltage-induced ATP synthesis by ATP-synthetases. In this case, F₀ is treated as a voltage modulating subunit whose function is to effect a periodic attenuation of the local electric field. This modulation allows multiple enzyme turnover even when a d.c. field is imposed. For the in vivo ATP synthesis, the membrane potential (d.c. in nature) presumably is supplied by the electron transport reactions. Reduction of cytochromes reconstituted into the lipid bilayer is well known to generate transmembrane potentials.The model treats the energy coupling event as a field-induced conformational change, and reduces energy transduction to events pertinent to the study of molecular mechanisms of enzyme action. The equations derived are solidly based on fundamental thermodynamic principles.