Studies on interactions between metmyoglobin and heparin.
The complex formation between metmyoglobin and heparin was investigated by absorbance and fluorescence spectroscopy as well as differential scanning microcalorimetry. In acidic pH region, three distinct complexes detected by absorbance measurements are formed depending on pH and time of equilibration. The kinetics of the conformational transition of metmyoglobin-heparin complex equilibrated at neutral pH observed after pH change to acidic region comprises two steps. During the first step, characterized by rapid changes of the absorption spectra (approximately 5 minutes) as well as fluorescence intensities, reversible transition with pK = 6.5 +/- 0.1 occurs and the first type of the complex forms. Below pH 6.2 the transition with pK = 5.7 +/- 0.1 is observed and the second type of the complex is formed. During the second slow step, the third type of the complex formed after 30 minutes of equilibration is characterized by a spectrum corresponding to low-spin form without protein axial ligand bound. At neutral pH and 25 degrees C, the interaction between metMb and heparin only slightly alters absorption and fluorescence spectra. On the other hand, the formation of metMb-heparin complex is established from the decrease of the transition temperature from 80.4 +/- 0.5 degrees C to 74.7 +/- 0.5 degrees C. Moreover, the binding of heparin prevents the aggregation of the protein at isoelectric point resulting in a considerable increase in the reversibility of thermal denaturation.
Department of Biophysics, Slovak Academy of Sciences, Kosice, Slovakia.
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Fedunová
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Antalík
D000818:Animals; D055592:Biophysical Phenomena; D001703:Biophysics; D002152:Calorimetry, Differential Scanning; D006493:Heparin / Q000737:chemistry / Q000378:metabolism*; D006736:Horses; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D046911:Macromolecular Substances; D008786:Metmyoglobin / Q000737:chemistry / Q000378:metabolism*; D011487:Protein Conformation; D011489:Protein Denaturation; D013050:Spectrometry, Fluorescence; D013053:Spectrophotometry; D013816:Thermodynamics; D014907:Whales
D046911:Macromolecular Substances; D008786:Metmyoglobin; D006493:Heparin