Records |
Author |
Abbruzzetti, S.; Viappiani, C.; Small, J.R.; Libertini, L.J.; Small, E.W. |
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 |
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 |
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Original Title |
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Series Title |
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Abbreviated Series Title |
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Series Volume |
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Series Issue |
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Edition |
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ISSN |
0002-7863 |
ISBN |
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Notes |
PMID:11439052 |
Approved |
no |
Call Number |
Equine Behaviour @ team @ |
Serial |
3788 |
Permanent link to this record |
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Author |
Steinhoff, H.J. |
Title |
A continuous wave laser T-jump apparatus and its application to chemical reactions in hemoglobin single crystals |
Type |
Journal Article |
Year |
1988 |
Publication |
Journal of Biochemical and Biophysical Methods |
Abbreviated Journal |
J Biochem Biophys Methods |
Volume |
15 |
Issue |
6 |
Pages |
319-330 |
Keywords |
Animals; Chemistry; Crystallization; *Heat; *Hemoglobins; Horses/blood; *Lasers; Methemoglobin; Solutions; Thermodynamics; Thiocyanates |
Abstract |
A laser temperature jump apparatus is constructed where the T-jump is achieved by means of the direct absorption of continuous laser radiation of low intensity by a solid sample. The final temperature in the irradiated volume element is reached when the absorbed radiation power equals the dissipation of heat by heat conduction. The time range from the beginning of irradiation to the stationary state depends on the geometry of the irradiated volume element and is less than 10 ms. The heating laser beam is simultaneously used to detect the relaxation to the new chemical equilibrium in the sample. Relaxation processes with relaxation rates between 10(2) s-1 and less than 10(-3) s-1 on samples with volumes less than 10(-3) mm3 may be investigated using this T-jump method. One application of this method is the determination of reaction rates of ligand reactions in hemoglobin single crystals. Rate constants obtained for the reaction of thiocyanate with crystallized horse methemoglobin are presented. |
Address |
Institut fur Biophysik, Ruhr-Universitat Bochum, F.R.G |
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Language |
English |
Summary Language |
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Original Title |
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Series Editor |
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Series Title |
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Abbreviated Series Title |
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Series Volume |
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Series Issue |
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Edition |
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ISSN |
0165-022X |
ISBN |
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Medium |
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Area |
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Expedition |
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Conference |
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Notes |
PMID:3379245 |
Approved |
no |
Call Number |
Equine Behaviour @ team @ |
Serial |
3804 |
Permanent link to this record |