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Author Bayley, P.; Martin, S.; Anson, M.
Title Temperature-jump circular dichroism: observation of chiroptical relaxation processes at millisecond time resolution Type Journal Article
Year (up) 1975 Publication Biochemical and Biophysical Research Communications Abbreviated Journal Biochem Biophys Res Commun
Volume 66 Issue 1 Pages 303-308
Keywords *Alcohol Oxidoreductases/metabolism; Animals; Circular Dichroism; Horses; Kinetics; Liver/enzymology; Mathematics; Protein Conformation; Temperature; Time Factors
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Language English Summary Language Original Title
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ISSN 0006-291X ISBN Medium
Area Expedition Conference
Notes PMID:1172440 Approved no
Call Number Equine Behaviour @ team @ Serial 3816
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Author Rodier, F.
Title [Spectral properties of porcine plasminogen: study of the acidic transition (author's transl)] Type Journal Article
Year (up) 1976 Publication European journal of biochemistry / FEBS Abbreviated Journal Eur J Biochem
Volume 63 Issue 2 Pages 553-562
Keywords Animals; Binding Sites; Guanidines; Hydrogen-Ion Concentration; *Plasminogen; Protein Binding; Protein Conformation; Spectrometry, Fluorescence; Spectrophotometry; Spectrophotometry, Ultraviolet; Swine; Temperature
Abstract The acidic transition of porcine plasminogen, prepared by affinity chromatography, was studied by non-destructive methods. These methods are based on the analysis of the behaviour of the tryptophyls under various conditions. The perturbation of the absorption and emission spectra by pH or temperature and the dynamic quenching of the intrinsic fluorescence are used to obtain information on structural changes which affect the environment of these residues. It is shown that by decreasing pH the fluorescence emission spectra are shifted toward the long wavelengths, with a broadening of the fluorescence band. The same effect can be obtained at constant pH by heating the protein solution. In order to analyze these phenomena, it is assumed that the fluorescence intensities at 355 nm and 328 nm reflect the proportion of the tryptophans which are exposed to the solvent, and buried, respectively. The plot of the ratio of the fluorescence intensities at these wavelengths versus pH or temperature leads to a titration curve showing an unmasking of tryptophans. The proportion of exposed tryptophans is measured by the dynamic fluorescence quenching technique and the data analyzed according to Lehrer. The plot of the fraction of exposed tryptophyls versus pH also shows the unmasking of these chromophores. Thermal perturbation of a solution of plaminogen at neutral pH induces a difference absorption spectrum whose amplitudes at the maxima are proportional to the number of exposed aromatic residues. The comparison with a solution of fully denatured plasminogen in 6 M guanidium chloride, where all the tryptophyls are exposed, shows that the percentage of exposure is equal to 59%. This number is significantly higher than the percentage found by the fluorescence quenching technique (20%), indicating that some tryptophyls are located in crevices, exposed to the solvent but not to the iodide. At acidic pH the absorption difference spectra induced by thermal perturbation are not classical, since they show an inversion and a new band between 300 nm and 305 nm. This band is mentioned in the literature as a minor band of tryptophan which appears when this chromophore is located in an asymmetric environment. On plotting the maximum amplitude of these spectra obtained at acidic pH versus temperature, we obtain a curve indicating that two types of antagonistic interactions are involved in the perturbation of the chromophores spectra. The spectrophotometric titration of plasminogen gives classical absorption difference spectra. By plotting the maximum amplitude at 292 nm versus pH, we obtain a titration curve with an apparent pK of 2.9 units. This pK is acidic which respect to the pK value of a normal carboxyl. This low value can be due to a positively charged group in the neighbourhood of a carboxyl, which interacts with one or more chromophores. When the carboxyl becomes protonated, this positively charged group is free and available to perturb the environment of some chromophores...
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Publisher Place of Publication Editor
Language French Summary Language Original Title Proprietes spectrales du plasminogene porcin. Etude de la transition acide
Series Editor Series Title Abbreviated Series Title
Series Volume Series Issue Edition
ISSN 0014-2956 ISBN Medium
Area Expedition Conference
Notes PMID:4326 Approved no
Call Number Admin @ knut @ Serial 22
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Author Tsong, T.Y.
Title Conformational relaxations of urea- and guanidine hydrochloride-unfolded ferricytochrome c Type Journal Article
Year (up) 1977 Publication The Journal of Biological Chemistry Abbreviated Journal J Biol Chem
Volume 252 Issue 24 Pages 8778-8780
Keywords *Cytochrome c Group; Guanidines/*pharmacology; Protein Conformation/drug effects; Spectrometry, Fluorescence; Urea/*pharmacology
Abstract Several recent studies of protein the unfolded proteins. In urea- and guanidine HCl-unfolded ferricytochrome c (horse heart), an acid-induced spin state transformation of the heme group has been detected by the heme absorptions, Trp-59 fluorescence, and the intrinsic viscosity of protein. Kinetics of this second conformational transition, by the temperature jump and stopped flow methods, are complex. One rapid reaction (tau1), pH-independent, occurs in a 50-mus range; the second reaction (tau2), in a 1-ms range, depends linearly upon pH and is faster at the alkaline side; a third reaction (tau3), in a 1-s range, shows a sigmoidal transition at pH 5.1 and is faster at the acidic side. The results are consistent with a kinetic scheme which involves protein conformational changes in the transformation of the heme coordination state. The kinetics, along with previous equilibrium studies, indicate that ligand or charge interactions within a protein molecule are not completely prohibited even in strongly denaturing conditions, such as in high concentrations of urea and guanidine HCl. Thus, local structures of peptide chain associated with these interactions can exist in the unfolded protein.
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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:200618 Approved no
Call Number refbase @ user @ Serial 3882
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Author Ridge, J.A.; Baldwin, R.L.; Labhardt, A.M.
Title Nature of the fast and slow refolding reactions of iron(III) cytochrome c Type Journal Article
Year (up) 1981 Publication Biochemistry Abbreviated Journal Biochemistry
Volume 20 Issue 6 Pages 1622-1630
Keywords Animals; Ascorbic Acid; *Cytochrome c Group; Guanidines; Horses; Kinetics; Oxidation-Reduction; Protein Conformation; Spectrum Analysis
Abstract The fast and slow refolding reactions of iron(III) cytochrome c (Fe(III) cyt c), previously studied by Ikai et al. (Ikai, A., Fish, W. W., & Tanford, C. (1973) J. Mol. Biol. 73, 165--184), have been reinvestigated. The fast reaction has the major amplitude (78%) and is 100-fold faster than the slow reaction in these conditions (pH 7.2, 25 degrees C, 1.75 M guanidine hydrochloride). We show here that native cyt c is the product formed in the fast reaction as well as in the slow reaction. Two probes have been used to test for formation of native cyt c. absorbance in the 695-nm band and rate of reduction of by L-ascorbate. Different unfolded species (UF, US) give rise to the fast and slow refolding reactions, as shown both by refolding assays at different times after unfolding (“double-jump” experiments) and by the formation of native cyt c in each of the fast and slow refolding reactions. Thus the fast refolding reaction is UF leads to N and the slow refolding reaction is Us leads to N, where N is native cyt c, and there is a US in equilibrium UF equilibrium in unfolded cyt c. The results are consistent with the UF in equilibrium US reaction being proline isomerization, but this has not yet been tested in detail. Folding intermediates have been detected in both reactions. In the UF leads to N reaction, the Soret absorbance change precedes the recovery of the native 695-nm band spectrum, showing that Soret absorbance monitors the formation of a folding intermediate. In the US leads to N reaction an ascorbate-reducible intermediate has been found at an early stage in folding and the Soret absorbance change occurs together with the change at 695 nm as N is formed in the final stage of folding.
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Language English Summary Language Original Title
Series Editor Series Title Abbreviated Series Title
Series Volume Series Issue Edition
ISSN 0006-2960 ISBN Medium
Area Expedition Conference
Notes PMID:6261802 Approved no
Call Number Equine Behaviour @ team @ Serial 3809
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Author Dyson, H.J.; Beattie, J.K.
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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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 Cho, K.C.; Chan, K.K.
Title Kinetics of cold-induced denaturation of metmyoglobin Type Journal Article
Year (up) 1984 Publication Biochimica et Biophysica Acta (BBA) – Protein Structure and Molecular Enzymology Abbreviated Journal
Volume 786 Issue 1-2 Pages 103-108
Keywords Metmyoglobin denaturation; Temperature jump; Denaturation kinetics; Conformational transformation; (Horse heart)
Abstract Using a slow temperature-jump spectrophotometer, we have studied the kinetics of cold-induced denaturation of metmyoglobin between 0[degree sign]C and 20[degree sign]C at acidic pH. The time-scale of the transition is slow and is of the order of minutes. The results are consistent with the transition's involving a total of three states, native (N), transient intermediate (I) and denatured (D), which are converted from one to the other in that order.
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Notes Approved no
Call Number refbase @ user @ Serial 3978
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Author Steinhoff, H.J.; Lieutenant, K.; Redhardt, A.
Title Conformational transition of aquomethemoglobin: intramolecular histidine E7 binding reaction to the heme iron in the temperature range between 220 K and 295 K as seen by EPR and temperature-jump measurements Type Journal Article
Year (up) 1989 Publication Biochimica et Biophysica Acta Abbreviated Journal Biochim Biophys Acta
Volume 996 Issue 1-2 Pages 49-56
Keywords Animals; Electron Spin Resonance Spectroscopy; Heme; Histidine; Horses; Humans; Hydrogen-Ion Concentration; Methemoglobin/*ultrastructure; Motion; Protein Conformation; Temperature; Thermodynamics; Water
Abstract Temperature-dependent EPR and temperature-jump measurements have been carried out, in order to examine the high-spin to low-spin transition of aquomethemogobin (pH 6.0). Relaxation rates and equilibrium constants could be determined as a function of temperature. As a reaction mechanism for the high-spin to low-spin transition, the binding of N epsilon of His E7 to the heme iron had been proposed; the same mechanism had been suggested for the ms-effect, found in temperature-jump experiments on aquomethemoglobin. A comparison of the thermodynamic quantities, deduced form the measurements in this paper, gives evidence that indeed the same reaction is investigated in both cases. Our results and most of the findings of earlier studies on the spin-state transitions of aquomethemoglobin, using susceptibility, optical, or EPR measurements, can be explained by the transition of methemoglobin with H2O as ligand (with high-spin state at all temperatures) and methemoglobin with ligand N epsilon of His E7 (with a low-spin ground state). Thermal fluctuations of large amplitude have to be postulated for the reaction to take place, so this reaction may be understood as a probe for the study of protein dynamics.
Address Institut fur Biophysik, Ruhr-Universitat Bochum, F.R.G
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ISSN 0006-3002 ISBN Medium
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Notes PMID:2544230 Approved no
Call Number Equine Behaviour @ team @ Serial 3803
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Author Koenen, E.P.C.; van Veldhuizen, A.E.; Brascamp, E.W.
Title Genetic parameters of linear scored conformation traits and their relation to dressage and show-jumping performance in the Dutch Warmblood Riding Horse population Type Journal Article
Year (up) 1995 Publication Livestock Production Science Abbreviated Journal
Volume 43 Issue 1 Pages 85-94
Keywords Horse; Heritability; Conformation; Dressage; Show jumping
Abstract In this study genetic parameters of linear scored conformation traits of the Dutch Warmblood Riding Horse were estimated in relation to performance in competition. Observations on 10 665 mares were analyzed with an animal model including the fixed effects age, classifier, location and percentage of thoroughbred. Using restricted maximum likelihood algorithms, heritabilities of 26 linear scored conformation traits were estimated in the range 0.09-0.28. Several conformation traits had high up to very high mutual genetic correlations. Competition results of 3476 horses with performance in dressage and 3220 horses with performance in show-jumping were linked to the conformation data to estimate the genetic relationship between conformation and performance in competition. The model for the evaluation of the competition results included the fixed effects riding club, age and sex. Estimated heritabilities for dressage and show-jumping were 0.17 +/- 0.05 and 0.19 +/- 0.04, respectively. Genetic correlations between conformation and performance were low to moderate. The length of the neck, length and position of the shoulders, shape and length of croup and muscularity of the haunches had a significant moderate genetic correlation with dressage. Muscularity of the neck, shape of the croup and muscularity of the haunches had a significant genetic correlation with show-jumping. The results indicate that, due to the low genetic correlations with performance traits, indirect selection for performance using conformation results is of limited value.
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Notes Approved no
Call Number refbase @ user @ Serial 3961
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Author Hagen, S.J.; Eaton, W.A.
Title Two-state expansion and collapse of a polypeptide Type Journal Article
Year (up) 2000 Publication Journal of Molecular Biology Abbreviated Journal J Mol Biol
Volume 301 Issue 4 Pages 1019-1027
Keywords Animals; Computer Simulation; Cytochrome c Group/*chemistry/*metabolism; Horses; Kinetics; Lasers; Models, Chemical; Peptides/*chemistry/*metabolism; Protein Conformation; Protein Denaturation; *Protein Folding; Spectrometry, Fluorescence; Temperature; Thermodynamics
Abstract The initial phase of folding for many proteins is presumed to be the collapse of the polypeptide chain from expanded to compact, but still denatured, conformations. Theory and simulations suggest that this collapse may be a two-state transition, characterized by barrier-crossing kinetics, while the collapse of homopolymers is continuous and multi-phasic. We have used a laser temperature-jump with fluorescence spectroscopy to measure the complete time-course of the collapse of denatured cytochrome c with nanosecond time resolution. We find the process to be exponential in time and thermally activated, with an apparent activation energy approximately 9 k(B)T (after correction for solvent viscosity). These results indicate that polypeptide collapse is kinetically a two-state transition. Because of the observed free energy barrier, the time scale of polypeptide collapse is dramatically slower than is predicted by Langevin models for homopolymer collapse.
Address Laboratory of Chemical Physics, NIDDK, National Institutes of Health, Building 5, Bethesda, MD, 20892-0520, USA
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Language English Summary Language Original Title
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ISSN 0022-2836 ISBN Medium
Area Expedition Conference
Notes PMID:10966803 Approved no
Call Number Equine Behaviour @ team @ Serial 3790
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Author Abbruzzetti, S.; Crema, E.; Masino, L.; Vecli, A.; Viappiani, C.; Small, J.R.; Libertini, L.J.; Small, E.W.
Title Fast events in protein folding: structural volume changes accompanying the early events in the N-->I transition of apomyoglobin induced by ultrafast pH jump Type Journal Article
Year (up) 2000 Publication Biophysical Journal Abbreviated Journal Biophys J
Volume 78 Issue 1 Pages 405-415
Keywords Animals; Apoproteins/*chemistry; Horses; *Hydrogen-Ion Concentration; Kinetics; Models, Molecular; Myoglobin/*chemistry; Protein Conformation; *Protein Folding; Protein Structure, Secondary; Spectrometry, Fluorescence
Abstract Ultrafast, laser-induced pH jump with time-resolved photoacoustic detection has been used to investigate the early protonation steps leading to the formation of the compact acid intermediate (I) of apomyoglobin (ApoMb). When ApoMb is in its native state (N) at pH 7.0, rapid acidification induced by a laser pulse leads to two parallel protonation processes. One reaction can be attributed to the binding of protons to the imidazole rings of His24 and His119. Reaction with imidazole leads to an unusually large contraction of -82 +/- 3 ml/mol, an enthalpy change of 8 +/- 1 kcal/mol, and an apparent bimolecular rate constant of (0.77 +/- 0.03) x 10(10) M(-1) s(-1). Our experiments evidence a rate-limiting step for this process at high ApoMb concentrations, characterized by a value of (0. 60 +/- 0.07) x 10(6) s(-1). The second protonation reaction at pH 7. 0 can be attributed to neutralization of carboxylate groups and is accompanied by an apparent expansion of 3.4 +/- 0.2 ml/mol, occurring with an apparent bimolecular rate constant of (1.25 +/- 0.02) x 10(11) M(-1) s(-1), and a reaction enthalpy of about 2 kcal/mol. The activation energy for the processes associated with the protonation of His24 and His119 is 16.2 +/- 0.9 kcal/mol, whereas that for the neutralization of carboxylates is 9.2 +/- 0.9 kcal/mol. At pH 4.5 ApoMb is in a partially unfolded state (I) and rapid acidification experiments evidence only the process assigned to carboxylate protonation. The unusually large contraction and the high energetic barrier observed at pH 7.0 for the protonation of the His residues suggests that the formation of the compact acid intermediate involves a rate-limiting step after protonation.
Address Dipartimento di Fisica, Universita di Parma, 43100 Parma, Italia
Corporate Author Thesis
Publisher Place of Publication Editor
Language English Summary Language Original Title
Series Editor Series Title Abbreviated Series Title
Series Volume Series Issue Edition
ISSN 0006-3495 ISBN Medium
Area Expedition Conference
Notes PMID:10620304 Approved no
Call Number Equine Behaviour @ team @ Serial 3792
Permanent link to this record