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H/D Exchange (d + exchange)
Terms modified by H/D Exchange Selected AbstractsSupramolecular encapsulation of 1,3-bis(1-adamantyl)imidazolium chloride by ,-cyclodextrins: towards inhibition of C(2)-H/D exchangeJOURNAL OF PHYSICAL ORGANIC CHEMISTRY, Issue 2 2009Loïc Leclercq Abstract The study of the hydrogen/deuterium exchange reactions of the C(2)-proton for different carbene precursors has been carried out in the absence and presence of ,-cyclodextrin in D2O at 25°C. Formation of the inclusion complexes of imidazolium salts with the native ,-cyclodextrin and the ,-dimethylcyclodextrin is demonstrated by 1D and 2D 1H NMR, ESI/HRMS and a molecular modelling study. Formation of the inclusion complexes of imidazolium salts with the native ,-cyclodextrin and the ,-dimethylcyclodextrin is a simple and efficient method to modify the acidity of the imidazolium H(2) and to modify its environment. Encapsulation of 1,3-disubstituted imidazolium chloride by ,-cyclodextrins results in the inhibition of the H(2)/D exchange in the complex. Copyright © 2008 John Wiley & Sons, Ltd. [source] Changes in protein conformation and dynamics upon complex formation of brain-derived neurotrophic factor and its receptor: Investigation by isotope-edited Fourier transform IR spectroscopyBIOPOLYMERS, Issue 1 2002Tiansheng Li Abstract The interactions of brain-derived neurotrophic factor (BDNF) with the extracellular domain of its receptor (trkB) are investigated by employing isotope-edited Fourier transform IR (FTIR) spectroscopy. The protein secondary structures of individual BDNF and trkB in solutions are compared with those in their complex. The temperature dependence of the secondary structures of BDNF, trkB, and their complex is also investigated. Consistent with the crystal structure, we observe by FTIR spectroscopy that BDNF in solution contains predominantly , strands (,53%) and relatively low contents of other secondary structures including , turns (,16%), disordered structures (,12%), and loops (,18%) and is deficient in , helix. We also observe that trkB in solution contains mostly , strands (52%) and little , helix. Conformational changes in both BDNF and trkB are observed upon complex formation. Specifically, upon binding of BDNF, the conformational changes in trkB appear to involve mostly , turns and disordered structures while the majority of the ,-strand conformation remains unchanged. The IR data indicate that some of the disordered structures in the loop regions are likely converted to , strands upon complex formation. The FTIR spectral data of BDNF, trkB, and their complex indicate that more amide NH groups of trkB undergo H,D exchange within the complex than those of the ligand-free receptor and that the thermal stability of trkB is decreased slightly upon binding of BDNF. The FT-Raman spectra of BDNF, trkB, and their complex show that the six intramolecular disulfide bonds of trkB undergo significant conformational changes upon binding of BDNF as a result of changes in the tertiary structure of trkB. Taken together, the FTIR and Raman data are consistent with the loosening of the tertiary structure of trkB upon binding of BDNF, which leads to more solvent exposure of the amide NH group and decreased thermal stability of trkB. This finding reveals an intriguing structural property of the neurotrophin ligand,receptor complex that is in contrast to other ligand,receptor complexes such as a cytokine,receptor complex that usually shows protection of the amide NH group and increased thermal stability upon complex formation. © 2002 John Wiley & Sons, Inc. Biopolymers (Biospectroscopy) 67: 10,19, 2002; DOI 10.1002/bip.10038 [source] Transition-Metal-Free Synthesis of Perdeuterated Imidazolium Ionic Liquids by Alkylation and H/D ExchangeEUROPEAN JOURNAL OF ORGANIC CHEMISTRY, Issue 17 2008Ralf Giernoth Abstract Economic, transition-metal-free syntheses of partially or completely deuterated imidazolium ionic liquids (ILs) were developed. Double alkylation starting from imidazole afforded side-chain deuterated imidazolium ionic liquids, which subsequently were fully deuterated by H/D-exchange on the cation ring. Isotopic exchange was studied for a range of ionic liquids, solvents and bases. Here, the presence of small amounts of basic impurities was found to significantly affect the exchange behaviour.(© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2008) [source] Platinum and Palladium Complexes Containing Cationic Ligands as Catalysts for Arene H/D Exchange and Oxidation,ANGEWANDTE CHEMIE, Issue 34 2010Marion Kationische Komplexe aus Palladium(II) oder Platin(II) und Bipyridinliganden mit Pyridiniumsubstituenten sind hoch aktive und beständige Katalysatoren für den H-D-Austausch und die Oxidation der C-H-Bindungen von aromatischen Substraten mit Umsatzzahlen bis 3200 und Umsatzfrequenzen bis 0.1,s,1. [source] Amino-phosphanes in RhI -Catalyzed Hydroformylation: New Mechanistic Insights Using D2O as Deuterium-Labeling AgentEUROPEAN JOURNAL OF INORGANIC CHEMISTRY, Issue 1 2006Jacques Andrieu Abstract In previous work, we have demonstrated that the dangling amino group in amino-phosphane ligands increases the rate of Rh-catalyzed styrene hydroformylation as a function of the amino group basicity and of the distance between the P and N functions. We now report additional stereochemical and mechanistic insights resulting from new catalytic experiments performed with Rh-,-P,N catalytic systems in the presence of D2O. In addition to the expected D0 product, the formation of the ,-D1 aldehyde, PhCH(CH2D)CHO was observed in all cases by 1H and 13C NMR spectroscopy, indicating that H/D exchange occurs for the rhodium-hydride complex. Minor amounts of a ,-D2 product, PhCH(CHD2)CHO, were also formed under certain conditions, demonstrating the reversibility of the olefin coordination step. The composition of the aldehyde mixture is slightly affected by the nature of the catalytic precursor or the P,N ligand used. In the specific case of the ,-P,N ligand [,-P,N = (SAr,SC)-Ph2PCH{o -C6H4Cl(Cr(CO)3)}NHPh], in combination with the [RhCl(COD)]2 precatalyst, products PhCD(CH3)CHO (,-D1) and PhCD(CH2D)CHO (,,,-D2) were also produced. This result suggests a reversible deprotonation assisted by an intramolecular H-bonding interaction between the dangling ammonium function and the carbonyl moiety. This isotopic exchange process decreases the asymmetric induction from 14 to 7,% ee when using the enantiopure version of this ligand. Aldehydes bearing a D atom on the formyl group, e.g. PhCH(CH3)CDO, were never observed. The latter observation excludes protonolysis of the rhodium-acyl intermediate as the aldehyde forming step. In addition, it also excludes a bimolecular reaction involving the rhodium-acyl and rhodium-hydride intermediates.(© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2006) [source] Hydrogen/Deuterium Exchange Reactions of Olefins with Deuterium Oxide Mediated by the Carbonylchlorohydrido- tris(triphenylphosphine)ruthenium(II) ComplexADVANCED SYNTHESIS & CATALYSIS (PREVIOUSLY: JOURNAL FUER PRAKTISCHE CHEMIE), Issue 9 2010Sunny Kai San Tse Abstract The catalytic properties of several ruthenium, osmium and rhodium hydride complexes for hydrogen/deuterium (H/D) exchange between olefins and deuterium oxide (D2O) were investigated. The most effective catalytic precursor was found to be the carbonylchlorohydridotris(triphenylphosphine)ruthenium(II) complex. Through H/D exchange between metal hydride and D2O, and reversible olefin insertion into an RuH(D) bond, protons attached to olefinic carbons and alkyl chains of olefins can all undergo H/D exchange with D2O. The catalytic reactions can be used to deuterate both terminal and internal olefins, for example, styrene, stilbene and cyclooctene. [source] Hydrogen/deuterium exchange kinetics of cytochrome C: An electrospray ionization fast flow experimentISRAEL JOURNAL OF CHEMISTRY, Issue 3-4 2003Orit Geller New experiments are described in which the gas phase hydrogen/deuterium (H/D) exchange kinetics is studied for multiply-protonated cytochrome c ions with +10 to +17 charges. The experimental technique involves electrospray ionization (ESI) combined with a fast-flow method. Experimental results are presented including (1) average rate constants for H/D exchange, (2) overall decay kinetics of the reactant ion, and (3) sets of profiles for consecutive deuterium exchanges as a function of the flow rate of ND3 as the deuterating agent. The maximum number of exchanged hydrogen atoms and the exchange rate are observed to increase with increasing charge. The +13 state demonstrates special reactivity with a reactant ion decay constant of 2.5 × 10,9 cc/molecule's. Further insight into the H/D exchange mechanism is anticipated upon analysis of the data with a newly developed algorithm for extracting site-specific rate constants from profiles for H/D exchange in gas phase protonated amino acids, their clusters, and peptides. The algorithm minimizes the mutual entropy or the Kullback-Leibler information divergence between the observed concentrations and a chosen model. [source] Convenient methods for the synthesis of d4, d2 and d6 isotopomers of 4-(4-fluorobenzyl)piperidineJOURNAL OF LABELLED COMPOUNDS AND RADIOPHARMACEUTICALS, Issue 6 2005Ágnes Proszenyák Abstract Pure 4-(4-fluoro-[2,3,5,6- 2H4]benzyl)piperidine was prepared via the Grignard reaction of 4-fluoro-[2,3,5,6- 2H4]bromobenzene and pyridine-4-aldehyde followed by consecutive deoxygenation and heteroatomic ring saturation in the presence of palladium on carbon catalyst. An improved method for the catalytic H/D exchange in benzylic positions of 4-(4-fluorobenzyl)piperidine and its d4 derivative has also been described. Copyright © 2005 John Wiley & Sons, Ltd. [source] Structure investigation of Maltacine C1a, C1b, C2a and C2b,cyclic peptide lactones of the Maltacine complex from Bacillus subtilisJOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 10 2005Gunnar Hagelin Abstract A new complex of cyclic peptide lactone antibiotics from Bacillus subtilis, which we named Maltacines, has recently been described. The structure elucidation of four of them is reported in this paper. The amino acid sequences and structures of the peptides were found by MSn of the ring-opened linear peptides, which gave uninterrupted sequences of Bn and Y,n ions. The identities of three unknown residues in the sequences were solved by a combination of derivatisation with phenylisothiocyanate (PITC), high-resolution mass spectrometry and H/D exchange. The nature and position of the cyclic structure was disclosed by a chemo-selective ring opening with Na18OH and was found to be a lactone formed between a hydroxyl of residue number 4 and the C -terminal amino acid number 12. For verification of the structure of the B2+ ion, peptides with different combinations of P/Q and P/K at the N -terminus were synthesised. The structure of the four peptides were found to be: C1a and C2a: cyclo-4,12(P-Q-Y-Adip-V-E-T-Y-Orn-103-Y-I-OH) and C1b/C2b: cyclo-4,12(P-Q-Y-Adip-V-E-T-Y-K-103-Y-I-OH). Adip = aminodihydroxy pentanoic acid. Copyright © 2005 John Wiley & Sons, Ltd. [source] Structure investigation of Maltacine D1a, D1b and D1c,cyclic peptide lactones of the Maltacine complex from Bacillus subtilisJOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 10 2005Gunnar Hagelin Abstract A new complex of cyclic peptide lactone antibiotics from Bacillus subtilis, which we named Maltacines has recently been described. The structure elucidation of three of them is reported in this paper. The amino acid sequences and structures of the peptides were found by MSn of the ring-opened linear peptides that gave uninterrupted sequences of Bn and Y,n ions. The identities of four unknown residues in the sequences were solved by a combination of derivatisation with phenylisothiocyanate (PITC), high-resolution mass spectrometry and H/D exchange. The nature and position of the cyclic structure was disclosed by a chemo-selective ring opening with Na18OH and was found to be a lactone formed between a hydroxyl of residue number 4 and the C -terminal amino acid number 12. For verification of the structure of the B2+ ion, peptides with different combinations of P/Q and P/K at the N -terminus were synthesized. The structures of the four peptides is tentatively suggested to be: D1a: cyclo(4,12)-P-Q-Y-Adip-A-E-T-Y-Orn-HGly-Y-I-OH, D1b: cyclo(4,12)-P-Q-Y-Adip-A-E-T-Y-Orn-S-Y-I-OH and D1c: cyclo(4,12)-P-Q-Y-Adip-A-E-T-Y-K-S-Y-I-OH. Adip = aminodihydroxy pentanoic acid and HGly = hydroxyglycine. Copyright © 2005 John Wiley & Sons, Ltd. [source] Isomer separation of hyperbranched polyesteramides with gas-phase H/D exchange and a novel MSn approach: DoDIPJOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 8 2002Sander Koster Abstract Two approaches are introduced that provide information about the isomeric composition of hyperbranched polyesteramides. The first approach is based on a novel tandem mass spectrometric (MSn) approach that allows the study of different types of isomeric structures by a separation based on their difference in appearance energy. The method is called DoDIP: dissociation of depleted ion populations. A first MS/MS step is used to fragment isomers with relatively low appearance energy. The isomers with higher appearance energy are fragmented in a second MS/MS step of higher energy. The second approach is based on gas-phase H/D exchange experiments that result in a bimodal isotopic distribution for oligomers XnDn+1 of which one distribution corresponds to a type of isomeric structure that exhibits H/D exchange behaviour and the other to an isomeric structure that does not exhibit H/D exchange behaviour. X is a difunctional anhydride of phthalic acid (P), 1,2-cyclohexanedicarboxylic acid (C), succinic acid (S) or glutaric acid (G). D in XnDn+1 is a trifunctional diisopropanolamine and n the degree of polymerization. The type of isomeric structure that does not exhibit H/D exchange behaviour has a non-alternating monomer sequence that contains an amine bond with a relatively high proton affinity. The other isomeric structure that does exhibit H/D exchange behaviour has an alternating monomer sequence containing only amide and ester bonds with relatively low proton affinity. Oligomer structures were confirmed with additional MS2 experiments after H/D exchange. H/D exchange experiments on the fragments obtained after MS2 of the parent ion show that next to previously postulated mechanisms for the cleavage of the ester and amide bond another reaction pathway must be operational. A new mechanism is introduced to explain the H/D exchange behaviour of the fragments that requires a cleavage of the amide bonds only. Two types of fragments are formed by this mechanism. One type is protonated due to the cleavage of the amide bond whereas the other type has an oxazolonium ion structure due to the loss of an additional H2O. Copyright © 2002 John Wiley & Sons, Ltd. [source] Mechanistic studies of amide bond scission during acidolytic deprotection of Pip containing peptideJOURNAL OF PEPTIDE SCIENCE, Issue 8 2008Chiara Rubini Abstract Unusual TFA catalyzed cleavage reaction is reported for peptide containing pipecolic acid residues. Although the use of TFA under standard cleavage conditions is sufficiently mild to prevent degradation of the desired products, the amide bond between consecutive pipecolic acid residues is unexpectedly hydrolyzed by standard TFA treatment. The hydrolysis is proposed to proceed via an oxazolinium ion intermediate. This mechanism is supported by H/D exchange as observed by ESI-MS and NMR experiments. Copyright © 2008 European Peptide Society and John Wiley & Sons, Ltd. [source] Supramolecular encapsulation of 1,3-bis(1-adamantyl)imidazolium chloride by ,-cyclodextrins: towards inhibition of C(2)-H/D exchangeJOURNAL OF PHYSICAL ORGANIC CHEMISTRY, Issue 2 2009Loïc Leclercq Abstract The study of the hydrogen/deuterium exchange reactions of the C(2)-proton for different carbene precursors has been carried out in the absence and presence of ,-cyclodextrin in D2O at 25°C. Formation of the inclusion complexes of imidazolium salts with the native ,-cyclodextrin and the ,-dimethylcyclodextrin is demonstrated by 1D and 2D 1H NMR, ESI/HRMS and a molecular modelling study. Formation of the inclusion complexes of imidazolium salts with the native ,-cyclodextrin and the ,-dimethylcyclodextrin is a simple and efficient method to modify the acidity of the imidazolium H(2) and to modify its environment. Encapsulation of 1,3-disubstituted imidazolium chloride by ,-cyclodextrins results in the inhibition of the H(2)/D exchange in the complex. Copyright © 2008 John Wiley & Sons, Ltd. [source] Reactivity of isobutane in fluorosulfonic based superacids,JOURNAL OF PHYSICAL ORGANIC CHEMISTRY, Issue 12 2002Alain Goeppert Abstract The behavior of isobutane in DSO3F containing various amounts of SbF5 has been studied in relation to the acid strength of the superacid system. In contrast to the DF,SbF5 system, H/D exchange occurs only in the weakest superacid via deprotonation of the t -butyl ion intermediate formed by an oxidative process. Kinetic isotope effect determination shows that the slow step is hydride transfer. At higher acidity, the increasing stability of this intermediate impedes isotopic exchange. Copyright © 2002 John Wiley & Sons, Ltd. [source] A multi-angular mass spectrometric view at cyclic nucleotide dependent protein kinases: In vivo characterization and structure/function relationshipsMASS SPECTROMETRY REVIEWS, Issue 4 2008Arjen Scholten Abstract Mass spectrometry has evolved in recent years to a well-accepted and increasingly important complementary technique in molecular and structural biology. Here we review the many contributions mass spectrometry based studies have made in recent years in our understanding of the important cyclic nucleotide activated protein kinase A (PKA) and protein kinase G (PKG). We both describe the characterization of kinase isozymes, substrate phosphorylation, binding partners and post-translational modifications by proteomics based methodologies as well as their structural and functional properties as revealed by native mass spectrometry, H/D exchange MS and ion mobility. Combining all these mass spectrometry based data with other biophysical and biochemical data has been of great help to unravel the intricate regulation of kinase function in the cell in all its magnificent complexity. © 2008 Wiley Periodicals, Inc. Mass Spec Rev 27: 331,353, 2008 [source] Thermodynamic stability measurements on multimeric proteins using a new H/D exchange- and matrix-assisted laser desorption/ionization (MALDI) mass spectrometry-based methodPROTEIN SCIENCE, Issue 4 2002Kendall D. Powell Abstract We recently reported on a new H/D exchange- and matrix-assisted laser desorption/ionization (MALDI) mass spectrometry-based technique, termed SUPREX, that removes several important limitations associated with measuring the thermodynamic stability of proteins. In contrast to conventional spectroscopy-based techniques for characterizing the equilibrium unfolding behavior of proteins, SUPREX is amenable to the thermodynamic analysis of both purified and unpurified proteins using mg to ng quantities of material. Here we report on the application of SUPREX to the analysis of multimeric protein systems. Included in this work are the SUPREX results we obtained in studies on six model multimeric proteins including the GCN4p1 dimer, the coil-VaLd trimer, the 4-oxalocrotonate tautomerase (4-OT) hexamer, the Trp repressor (TrpR) dimer, the Arc repressor (ArcR) dimer, and an ArcR mutant (the (DOA20)ArcR) dimer which contained two destabilizing mutations including an Asp to Ala mutation at position 20 and an amide to ester bond mutation between amino acid (aa) residues 19 and 20. As part of the work described here, we present a new method for the analysis of SUPREX data that is generally applicable to both monomeric and multimeric protein systems. Our results on the model proteins in this study indicate that this new method can be used to determine folding free energies for proteins with the accuracy and precision of conventional spectroscopy-based methods. [source] Direct evidence by H/D exchange and ESI-MS for transient unproductive domain interaction in the refolding of an antibody scFv fragmentPROTEIN SCIENCE, Issue 3 2000Marcus Jäger Abstract The refolding kinetics of a single-chain Fv (scFv) fragment, derived from a stabilized mutant of the phosphorylcholine binding antibody McPC603, was investigated by H/D exchange and ESI-MS and compared with the folding kinetics of its constituting domains VH and VL. Both VH and VL adopt essentially native-like exchange protection within the dead time of the manual-mixing H/D exchange experiment (10 s) and in the case of VL, which contains two cis -prolines in the native conformation, this fast protection is independent of proline cis/trans isomerization. At the earliest time point resolvable by manual mixing, fewer deuterons are protected in the scFv fragment than in the two isolated domains together, despite the fact that the scFv fragment is significantly more stable than VL and VH. Full H/D exchange protection in the scFv fragment is gained on a time scale of minutes. This means that the domains in the scFv fragment do not refold independently. Rather, they associate prematurely and in nonnative form, a kinetic trap. Unproductive domain association is observed both after equilibrium- and short-term denaturation. For the equilibrium-denatured scFv fragment, whose native structure formation is dependent on a cis conformation of an interface proline in VL, this cis/trans isomerization reaction proceeds about one order in magnitude more slowly than the escape from the trap to a conformation where full H/D exchange protection is already achieved. We interpret these data in terms of a general kinetic scheme involving intermediates with and without domain association. [source] Neutron protein crystallography: beyond the folding structure of biological macromoleculesACTA CRYSTALLOGRAPHICA SECTION A, Issue 1 2008Nobuo Niimura Neutron diffraction provides an experimental method of directly locating H atoms in proteins, a technique complementary to ultra-high-resolution X-ray diffraction. Three different types of neutron diffractometers for biological macromolecules have been constructed in Japan, France and the USA, and they have been used to determine the crystal structures of proteins up to resolution limits of 1.5,2.5,Å. Results relating to H-atom positions and hydration patterns in proteins have been obtained from these studies. Examples include the geometrical details of hydrogen bonds, the role of H atoms in enzymatic activity, CH3 configuration, H/D exchange in proteins and oligonucleotides, and the dynamical behavior of hydration structures, all of which have been extracted from these structural results and reviewed. Other techniques, such as the growth of large single crystals and a database of hydrogen and hydration in proteins, are described. [source] Mass spectrometric investigation of Maltacines E1a and E1b,two members of the Maltacine family of peptide antibioticsRAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 24 2005Gunnar Hagelin We have recently described the discovery of the Maltacines,a new family of cyclic peptide antibiotics from Bacillus subtilis. In this paper the mass spectrometric characterisation of two of the members is reported. A chemoselective ring opening with base to give the linear peptides was necessary before mass spectrometric characterisation could be performed. MSn of the singly and doubly charged protonated molecules gave uninterrupted series of Bn and Y,n ions that allowed determination of the amino acid sequence. By using a combination of derivatisation with phenylisothiocyanate (PITC), high-resolution mass spectrometry and H/D exchange, the identities of three unknown residues were determined. The nature and position of the cyclic structure were disclosed by a chemoselective ring opening with Na18OH and it was found to be a lactone formed between a hydroxyl of residue number 4 and the C-terminal amino acid number 12. Peptides with different combinations of P/Q and P/K at the N-terminus were synthesised to verify the sequence of the N-terminal B2 ion. The structure of the two peptides is proposed to be: E1a: cyclo-4,12(P-Q-Y-Adip-V-E-T-Y-Orn-S-Y-I-OH) and E1b: cyclo-4,12(P-Q-Y-Adip-V-E-T-Y-K-S-Y-I-OH). Adip,=,(2-amino-4,5-dihydroxypentanoic acid). Copyright © 2005 John Wiley & Sons, Ltd. [source] Denaturant sensitive regions in creatine kinase identified by hydrogen/deuterium exchangeRAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 11 2005Hortense Mazon The GdmHCl-induced unfolding of creatine kinase (CK) has been studied by hydrogen/deuterium (H/D) exchange combined with mass spectrometry. MM-CK unfolded for various periods in different denaturant concentrations was pulsed-labeled with deuterium to identify different conformational intermediate states. For all denaturation times or GdmHCl concentrations, we observed variable proportions of only two species. The low-mass envelope of isotope peaks corresponds to a species that has gained about 10 deuteriums more than native CK, and the high-mass envelope to a completely deuterated species. To localize precisely the unfolded regions in the states highly populated during denaturation, the protein was digested with two proteases (pepsin and type XIII protease) after H/D exchange and rapid quenching of the reaction. The two sets of fragments obtained were analyzed by liquid chromatography coupled to mass spectrometry to determine the deuterium level in each fragment. Bimodal distributions of deuterium were found for most peptides, indicating that these regions were either folded or unfolded. This behavior is consistent with cooperative, localized unfolding. However, we observed a monomodal distribution of deuterium in two regions (1,12 and 162,186). We conclude that the increment of mass observed in the low-mass species of the intact protein (+10,Da) has its origin in these two segments. These regions, which are very sensitive to low GdmHCl concentrations, are involved in the monomer,monomer interface of CK and their perturbation is likely to weaken the dimeric structure. At higher denaturant concentration, this would induce dissociation of the dimer. Copyright © 2005 John Wiley & Sons, Ltd. [source] A neutron crystallographic analysis of T6 porcine insulin at 2.1,Å resolutionACTA CRYSTALLOGRAPHICA SECTION D, Issue 10 2009Wakari Iwai Neutron diffraction data for T6 porcine insulin were collected to 2.1,Å resolution from a single crystal partly deuterated by exchange of mother liquor. A maximum-likelihood structure refinement was undertaken using the neutron data and the structure was refined to a residual of 0.179. The hydrogen-bonding network of the central core of the hexamer was observed and the charge balance between positively charged Zn ions and their surrounding structure was interpreted by considering the protonation and/or deprotonation states and interactions of HisB10, water and GluB13. The observed double conformation of GluB13 was essential to interpreting the charge balance and could be compared with the structure of a dried crystal of T6 human insulin at 100,K. Differences in the dynamic behaviour of the water molecules coordinating the upper and lower Zn ions were observed and interpreted. The hydrogen bonds in the insulin molecules, as well as those involving HisB10 and GluB13, are discussed. The hydrogen/deuterium (H/D) exchange ratios of the amide H atoms of T6 porcine insulin in crystals were obtained and showed that regions highly protected from H/D exchange are concentrated in the centre of a helical region of the B chains. From the viewpoint of soaking time versus H/D-exchange ratios, the amide H atoms can be classified into three categories. [source] Glycerol-induced folding of unstructured disulfide-deficient lysozyme into a native-like conformationBIOPOLYMERS, Issue 8 2009Keiko Sakamoto Abstract 2SS[6-127,64-80] variant of lysozyme which has two disulfide bridges, Cys6-Cys127 and Cys64-Cys80, and lacks the other two disulfide bridges, Cys30-Cys115 and Cys76-Cys94, was quite unstructured in water, but a part of the polypeptide chain was gradually frozen into a native-like conformation with increasing glycerol concentration. It was monitored from the protection factors of amide hydrogens against H/D exchange. In solution containing various concentrations of glycerol, H/D exchange reactions were carried out at pH* 3.0 and 4°C. Then, 1H- 15N-HSQC spectra of partially deuterated protein were measured in a quenching buffer for H/D exchange (95% DMSO/5% D2O mixture at pH* 5.5 adjusted with dichloroacetate). In a solution of 10% glycerol, the protection factors were nearly equal to 10 at most of residues. With increasing glycerol concentration, some selected regions were further protected, and their protection factors reached about a 1000 in 30% glycerol solution. The highly protected residues were included in A-, B-, and C-helices and ,3-strand, and especially centered on Ile 55 and Leu 56. In 2SS[6-127,64-80], long-range interactions were recovered due to the preferential hydration by glycerol in the hydrophobic box of the ,-domain. Glycerol-induced recovering of the native-like structure is discussed from the viewpoint of molten globules growing with the protein folding. © 2009 Wiley Periodicals, Inc. Biopolymers 91: 665,675, 2009. This article was originally published online as an accepted preprint. The "Published Online" date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com [source] Significant Influence of Zn on Activation of the C-H Bonds of Small Alkanes by Brønsted Acid Sites of Zeolite,CHEMPHYSCHEM, Issue 17 2008Alexander G. Stepanov Prof. Abstract Herein, we analyze earlier obtained and new data about peculiarities of the H/D hydrogen exchange of small C1,n -C4 alkanes on Zn-modified high-silica zeolites ZSM-5 and BEA in comparison with the exchange for corresponding purely acidic forms of these zeolites. This allows us to identify an evident promoting effect of Zn on the activation of CH bonds of alkanes by zeolite Brønsted sites. The effect of Zn is demonstrated by observing the regioselectivity of the H/D exchange for propane and n- butane as well as by the increase in the rate and a decrease in the apparent activation energy of the exchange for all C1,n -C4 alkanes upon modification of zeolites with Zn. The influence of Zn on alkane activation has been rationalized by dissociative adsorption of alkanes on Zn oxide species inside zeolite pores, which precedes the interaction of alkane with Brønsted acid sites. [source] H/D exchange reactions;CHEMPHYSCHEM, Issue 6 2003sigma-bond metathesis; The mechanism of the H/D exchange reaction in alkane/hydrogen mixtures on silica-supported zirconium hydride was investigated by a modelling study using density functional theory (DFT) calculations. The electronic activation enthalpy (,H) for the CH bond activation step (TS3) was calculated to be around 92 kJ,mol,1, whereas it would be 258 kJ,mol,1for a direct exchange process (TS1, also called the kite TS). These data clearly speak in favour of the former as a mechanism for CH bond scrambling. Moreover, the calculated enthalpy of activation (,H) for H/D exchange in H2/D2mixtures (TS2) is 33.5 kJ,mol,1, which shows that this reaction is much faster than the H/D scrambling in alkane/H2mixtures, as shown experimentally. Additionally, the calculated activation entropies (For TS1,4,,S ranges between ,129 and ,174 J,mol,1,K,1) are very negative. Although the calculated activation entropies are also in full agreement with experimental data (,S=,113 J,mol,1,K,1), overall, the calculated activation enthalpies are much higher than the experimental ones. This suggests that the actual catalyst is probably more electrophilic than the model chosen for the calculations. [source] Study of Structural Stability of Cyclophilin A by NMR and Circular Dichroism SpectraCHINESE JOURNAL OF CHEMISTRY, Issue 7 2006Yan-Hong Shi Abstract The structural stability of cyclophilin A (CypA) was investigated using H/D exchange and temperature coefficients of chemical shifts of amide protons, monitored by 2D heteronuclear NMR spectroscopy. Amide proton exchange rates were measured by H/D exchange experiments for slow-exchange protons and measured by SEA (Solvent Exposed Amides)-HSQC experiments for fast-exchange protons. Temperature coefficients of chemical shifts and hydrogen exchange rates of amide protons show reasonably good correlation with the protein structure. Totally, 44 out of 153 non-proline assigned residues still exist in 86 d of hydrogen-deuterium exchange at 4 °C, suggesting that CypA structure should be highly stable. Residues in secondary structures of ,2, ,1, ,2, ,5, ,6 and ,7 might constitute the hydrophobic core of the protein. The change in free energy of unfolding (,Gu) of CypA was estimated to be (21.99±1.53) kJ·mol,1 by circular dichroism (CD). The large free energy change is also an indicator of the high structural stability. [source] | |||||||||||||||||||||||||