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Author Andersson, P.; Kvassman, J.; Lindstrom, A.; Olden, B.; Pettersson, G. openurl 
  Title Effect of NADH on the pKa of zinc-bound water in liver alcohol dehydrogenase Type Journal Article
  Year (up) 1981 Publication European Journal of Biochemistry / FEBS Abbreviated Journal Eur J Biochem  
  Volume 113 Issue 3 Pages 425-433  
  Keywords Alcohol Oxidoreductases/*metabolism; Aldehydes/metabolism; Animals; Binding Sites; Cinnamates/metabolism; Horses; Hydrogen-Ion Concentration; Kinetics; Ligands; Liver/*metabolism; NAD/*metabolism; Water/metabolism; Zinc/metabolism  
  Abstract Equilibrium constants for coenzyme binding to liver alcohol dehydrogenase have been determined over the pH range 10--12 by pH-jump stop-flow techniques. The binding of NADH or NAD+ requires the protonated form of an ionizing group (distinct from zinc-bound water) with a pKa of 10.4. Complex formation with NADH exhibits an additional dependence on the protonation state of an ionizing group with a pKa of 11.2. The binding of trans-N,N-dimethylaminocinnamaldehyde to the enzyme . NADH complex is prevented by ionization of the latter group. It is concluded from these results that the pKa-11.2-dependence of NADH binding most likely derives from ionization of the water molecule bound at the catalytic zinc ion of the enzyme subunit. The pKa value of 11.2 thus assigned to zinc-bound water in the enzyme . NADH complex appears to be typical for an aquo ligand in the inner-sphere ligand field provided by the zinc-binding amino acid residues in liver alcohol dehydrogenase. This means that the pKa of metal-bound water in zinc-containing enzymes can be assumed to correlate primarily with the number of negatively charged protein ligands coordinated by the active-site zinc ion.  
  Address  
  Corporate Author Thesis  
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  Language English Summary Language Original Title  
  Series Editor Series Title Abbreviated Series Title  
  Series Volume Series Issue Edition  
  ISSN 0014-2956 ISBN Medium  
  Area Expedition Conference  
  Notes PMID:7011796 Approved no  
  Call Number Equine Behaviour @ team @ Serial 3810  
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Author Dyson, H.J.; Beattie, J.K. openurl 
  Title Spin state and unfolding equilibria of ferricytochrome c in acidic solutions Type Journal Article
  Year (up) 1982 Publication The Journal of Biological Chemistry Abbreviated Journal J Biol Chem  
  Volume 257 Issue 5 Pages 2267-2273  
  Keywords Animals; *Cytochrome c Group; Electron Spin Resonance Spectroscopy; Heme; Horses; Hydrogen-Ion Concentration; Kinetics; Ligands; Myocardium; Protein Binding; Protein Conformation; Spectrophotometry; Temperature  
  Abstract Equilibrium, stopped flow, and temperature-jump spectrophotometry have been used to identify processes in the unfolding of ferricytochrome c in acidic aqueous solutions. A relaxation occurring in approximately 100 microseconds involves perturbation of a spin-equilibrium between two folded conformers of the protein with methionine-80 coordinated or dissociated from the heme iron. The protein unfolds more slowly, in milliseconds, with dissociation and protonation of histidine-18. These two transitions appear cooperative in equilibrium measurements at low (0.01 M) ionic strength, but are separated at higher (0.10 M) ionic strength. They are resolved under both conditions in the dynamic measurements. The spin-equilibrium description permits a unified explanation of a number of properties of ferricytochrome c in acidic aqueous solutions.  
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  Corporate Author Thesis  
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  Language English Summary Language Original Title  
  Series Editor Series Title Abbreviated Series Title  
  Series Volume Series Issue Edition  
  ISSN 0021-9258 ISBN Medium  
  Area Expedition Conference  
  Notes PMID:6277891 Approved no  
  Call Number Equine Behaviour @ team @ Serial 3807  
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Author Abbruzzetti, S.; Viappiani, C.; Small, J.R.; Libertini, L.J.; Small, E.W. openurl 
  Title Kinetics of histidine deligation from the heme in GuHCl-unfolded Fe(III) cytochrome C studied by a laser-induced pH-jump technique Type Journal Article
  Year (up) 2001 Publication Journal of the American Chemical Society Abbreviated Journal J Am Chem Soc  
  Volume 123 Issue 27 Pages 6649-6653  
  Keywords Animals; *Bacterial Proteins; Cytochrome c Group/*chemistry; Guanidine/*chemistry; Heme/*chemistry; Histidine/*chemistry; Horses; Hydrogen-Ion Concentration; Kinetics; *Lasers; Ligands; Protein Folding  
  Abstract We have developed an instrumental setup that uses transient absorption to monitor protein folding/unfolding processes following a laser-induced, ultrafast release of protons from o-nitrobenzaldehyde. The resulting increase in [H(+)], which can be more than 100 microM, is complete within a few nanoseconds. The increase in [H(+)] lowers the pH of the solution from neutrality to approximately 4 at the highest laser pulse energy used. Protein structural rearrangements can be followed by transient absorption, with kinetic monitoring over a broad time range (approximately 10 ns to 500 ms). Using this pH-jump/transient absorption technique, we have examined the dissociation kinetics of non-native axial heme ligands (either histidine His26 or His33) in GuHCl-unfolded Fe(III) cytochrome c (cyt c). Deligation of the non-native ligands following the acidic pH-jump occurs as a biexponential process with different pre-exponential factors. The pre-exponential factors markedly depend on the extent of the pH-jump, as expected from differences in the pK(a) values of His26 and His33. The two lifetimes were found to depend on temperature but were not functions of either the magnitude of the pH-jump or the pre-pulse pH of the solution. The activation energies of the deligation processes support the suggestion that GuHCl-unfolded cyt c structures with non-native histidine axial ligands represent kinetic traps in unfolding.  
  Address Dipartimento di Fisica, Universita di Parma, Istituto Nazionale per la Fisica della Materia, 43100 Parma, Italy  
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  Language English Summary Language Original Title  
  Series Editor Series Title Abbreviated Series Title  
  Series Volume Series Issue Edition  
  ISSN 0002-7863 ISBN Medium  
  Area Expedition Conference  
  Notes PMID:11439052 Approved no  
  Call Number Equine Behaviour @ team @ Serial 3788  
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Author Abbruzzetti, S.; Viappiani, C.; Sinibaldi, F.; Santucci, R. openurl 
  Title Kinetics of histidine dissociation from the heme Fe(III) in N-fragment (residues 1-56) of cytochrome c Type Journal Article
  Year (up) 2004 Publication The Protein Journal Abbreviated Journal Protein J  
  Volume 23 Issue 8 Pages 519-527  
  Keywords Animals; Cytochromes c/*chemistry; Enzyme Activation; Histidine/*chemistry; Horses; Hydrogen-Ion Concentration; Kinetics; Lasers; Ligands; Peptide Mapping; Photolysis; Spectrophotometry  
  Abstract We have here investigated the dissociation kinetics of the His side chains axially ligated to the heme-iron in the ferric (1-56 residues) N-fragment of horse cyt c. The ligand deligation induced by acidic pH-jump occurs as a biexponential process with different pre-exponential factors, consistent with a structural heterogeneity in solution and the presence of two differently coordinated species. In analogy with GuHCl-denatured cyt c, our data indicate the presence in solution of two ferric forms of the N-fragment characterized by bis-His coordination, as summarized in the following scheme: His18-Fe(III)-His26 <==> His18-Fe(III)-His33. We have found that the pre-exponential factors depend on the extent of the pH-jump. This may be correlated with the different pKa values shown by His26 and His33; due to steric factors, His26 binds to the heme-Fe(III) less strongly than His33, as recently shown by studies on denatured cyt c. Interestingly, the two lifetimes are affected by temperature but not by the extent of the pH-jump. The lower pKa for the deligation reaction required the use of an improved laser pH-jump setup, capable of inducing changes in H+ concentration as large as 1 mM after the end of the laser pulse. For the ferric N-fragment, close activation entropy values have been determined for the two histidines coordinated to the iron; this result significantly differs from that for GuHCl-denatured cyt c, where largely different values of activation entropy were calculated. This underlines the role played by the missing segment (residues 57-104) peptide chain in discriminating deligation of the “nonnative” His from the sixth coordination position of the metal.  
  Address Dipartimento di Fisica, Universita degli Studi di Parma, Parco Area delle Scienze 7/A 43100 Parma, Italy  
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  Language English Summary Language Original Title  
  Series Editor Series Title Abbreviated Series Title  
  Series Volume Series Issue Edition  
  ISSN 1572-3887 ISBN Medium  
  Area Expedition Conference  
  Notes PMID:15648974 Approved no  
  Call Number Equine Behaviour @ team @ Serial 3770  
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Author Hirota, S.; Suzuki, M.; Watanabe, Y. openurl 
  Title Hydrophobic effect of trityrosine on heme ligand exchange during folding of cytochrome c Type Journal Article
  Year (up) 2004 Publication Biochemical and Biophysical Research Communications Abbreviated Journal Biochem Biophys Res Commun  
  Volume 314 Issue 2 Pages 452-458  
  Keywords Amino Acids/chemistry; Animals; Cytochromes c/*chemistry; Heme/*chemistry; Histidine/chemistry; Horses; Hydrogen-Ion Concentration; Kinetics; Ligands; Myocardium/chemistry; Peptides/chemistry; Protein Folding; Spectrophotometry; Spectrum Analysis, Raman; Tyrosine/*analogs & derivatives/*chemistry  
  Abstract Effect of a hydrophobic peptide on folding of oxidized cytochrome c (cyt c) is studied with trityrosine. Folding of cyt c was initiated by pH jump from 2.3 (acid-unfolded) to 4.2 (folded). The Soret band of the 2-ms transient absorption spectrum during folding decreased its intensity and red-shifted from 397 to 400 nm by interaction with trityrosine, whereas tyrosinol caused no significant effect. The change in the transient absorption spectrum by interaction with trityrosine was similar to that obtained with 100 mM imidazole, which showed that the population of the intermediate His/His coordinated species increased during folding of cyt c by interaction with trityrosine. The absorption change was biphasic, the fast phase (82+/-9s(-1)) corresponding to the transition from the His/H(2)O to the His/Met coordinated species, whereas the slow phase (24+/-3s(-1)) from His/His to His/Met. By addition of trityrosine, the relative ratio of the slow phase increased, due to increase of the His/His species at the initial stage of folding. According to the resonance Raman spectra of cyt c, the high-spin 6-coordinate and low-spin 6-coordinate species were dominated at pH 2.3 and 4.2, respectively, and these species were not affected by addition of trityrosine. These results demonstrated that the His/His species increased by interaction with trityrosine at the initial stage of cyt c folding, whereas the heme coordination structure was not affected by trityrosine when the protein was completely unfolded or folded. Hydrophobic peptides thus may be useful to study the effects of hydrophobic interactions on protein folding.  
  Address Department of Physical Chemistry, Kyoto Pharmaceutical University, Yamashina-ku, 607-8414 Kyoto, Japan. hirota@mb.kyoto-phu.ac.jp  
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  Language English Summary Language Original Title  
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  Series Volume Series Issue Edition  
  ISSN 0006-291X ISBN Medium  
  Area Expedition Conference  
  Notes PMID:14733927 Approved no  
  Call Number Equine Behaviour @ team @ Serial 3777  
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Author Nicol, C.J.; Adachi, M.; Akiyama, T.E.; Gonzalez, F.J. doi  openurl
  Title PPARgamma in endothelial cells influences high fat diet-induced hypertension Type Journal Article
  Year (up) 2005 Publication American journal of hypertension : journal of the American Society of Hypertension Abbreviated Journal Am J Hypertens  
  Volume 18 Issue 4 Pt 1 Pages 549-556  
  Keywords Administration, Oral; Animals; Antihypertensive Agents/pharmacology; Blood Pressure/drug effects; Diabetes Mellitus, Type 2/physiopathology; Dietary Fats/*administration & dosage/pharmacology; Dose-Response Relationship, Drug; Endothelial Cells/*metabolism; Female; Heart Rate/drug effects; Hypertension/*etiology; Ligands; Male; Mice; Mice, Knockout; PPAR gamma/*metabolism; Sodium Chloride/administration & dosage/pharmacology; Thiazolidinediones/pharmacology  
  Abstract BACKGROUND: Peroxisome proliferator-activated receptor gamma (PPARgamma) ligands improve human hypertension. However, the mechanism and site of this effect remains unknown, confounded by PPARgamma expression in many cell types, including endothelial cells (ECs). METHODS: To evaluate the vascular role of PPARgamma we used a conditional null mouse model. Specific disruption of PPARgamma in ECs was created by crossing Tie2-Cre+ transgenic (T2T+) and PPARgamma-floxed (fl/fl) mice to generate PPARgamma (fl/fl)T2T+ (PPARgamma E-null) mice. Conscious 8- to 12-week-old congenic PPARgamma (fl/fl)Cre- (wild type) and PPARgamma E-null mice were examined for changes in systolic blood pressure (BP) and heart rate (HR), untreated, after 2 months of salt-loading (drinking water), and after treatment for 3 months with high fat (HF) diet alone or supplemented during the last 2 weeks with rosiglitazone (3 mg/kg/d). RESULTS: Untreated PPARgamma E-nulls were phenotypically indistinguishable from wild-type littermates. However, compared to similarly treated wild types, HF-treated PPARgamma E-nulls had significantly elevated systolic BP not seen after normal diet or salt-loading. Despite sex-dependent baseline differences, salt-loaded and HF-treated PPARgamma E-nulls of either sex had significantly elevated HR versus wild types. Interestingly, rosiglitazone improved serum insulin levels, but not HF diet-induced hypertension, in PPARgamma E-null mice. CONCLUSIONS: These results suggest that PPARgamma in ECs not only is an important regulator of hypertension and HR under stressed conditions mimicking those arising in type 2 diabetics, but also mediates the antihypertensive effects of rosiglitazone. These data add evidence supporting a beneficial role for PPARgamma-specific ligands in the treatment of hypertension, and suggest therapeutic strategies targeting ECs may prove useful.  
  Address Laboratory of Metabolism, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA  
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  Language English Summary Language Original Title  
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  Series Volume Series Issue Edition  
  ISSN 0895-7061 ISBN Medium  
  Area Expedition Conference  
  Notes PMID:15831367 Approved no  
  Call Number refbase @ user @ Serial 69  
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