ABSTRACT
X-ray studies revealed the considerable enhancement of metal-binding properties in human hemoglobin under exposure to mild damaging factors (in the presence of 0.09 M urea or upon heating for 30 min at 50 °C). Changes in the element composition of the hemoglobin monolayer, formed on the water subphase in the Langmuir trough, have been monitored in real time by the total external reflection X-ray fluorescence measurements. X-ray absorption spectroscopy has been applied to study the local environment of zinc ions bound on hemoglobin molecules. According to these data, each zinc ion is coordinated by four ligands, two of which are cysteine and histidine. The oxidative stress has been found to accelerate extensively the enhancement of metal-binding ability in protein. A two-stage mechanism has been proposed as a possible explanation of the observed phenomenon: First, in the presence of the mild damaging agents, protein molecules can undergo a transition from the native conformation to a more labile intermediate state that increases the accessibility of amino acid residues (in particular cysteine). At the second stage, oxidation of cysteine and the subsequent activation of cysteine SH groups can affect markedly the protein-metal interaction. The presented investigations provide a deeper insight into the pathogenesis of metabolic disorders that excessive concentrations of the endogenic toxicants might trigger in an organism.
Subject(s)
Hemoglobins/chemistry , Hemoglobins/metabolism , Metals/metabolism , Humans , Models, Molecular , Protein Binding/drug effects , Protein Conformation , Surface Properties , Urea/pharmacologyABSTRACT
Proteins of the transferrin (Tf) family have a role in metal transport in vertebrates and have been extensively studied. The results here reported provide, for the first time, a detailed systematic comparison of metal sites in Tf complexes involving several atoms in the whole protein and in two different types of Tfs. The high interest in the structural variations induced in a metalloprotein upon the uptake of different metals is related to the hypothesis of the metals' involvement in some neuropathologies. We propose a comparative study of the X-ray absorption spectra at the K-edge of iron, copper, zinc and nickel in serotransferrin and ovotransferrin. The experimental data are simulated using an algorithm of the full multiple scattering method. Our results show that: (1) the local structure of each site (N-terminal and C-terminal) is correlated to the ligation state of the other site; (2) the difference between the two proteins is related to site local structure and depends on the metal ion nature being greater in the case of copper and zinc with respect to iron and nickel ions; (3) X-ray spectroscopy is confirmed as a suitable technique able to discriminate between coordination models proposed by X-ray diffraction.
Subject(s)
Conalbumin/chemistry , Metals/chemistry , Models, Molecular , Spectrometry, X-Ray Emission/methods , Transferrin/chemistry , Animals , Binding Sites , Chickens , Computer Simulation , Humans , Protein Binding , Protein Conformation , Species SpecificityABSTRACT
The active metal site structure of transferrin with iron and copper atoms is investigated using metal K-XANES. Theoretical analysis of experimental data has been performed on the basis of full multiple-scattering theory. This approach made it possible to study the origin of XANES fine details and to investigate the local structure around active metal sites. A deep insight into the local structure and electronic subsystem of Fe, Cu transferrins is obtained. For example, in the case of Cu substitution of Fe in the active centre, the best fit of theoretical spectra to experiment has been obtained for distances 3% smaller between the Cu atom and the nearest neighbours.
Subject(s)
Conalbumin/chemistry , Copper/chemistry , Iron/chemistry , Transferrin/chemistry , Animals , Binding Sites , Chickens , Female , Humans , Spectrometry, X-Ray Emission/methodsABSTRACT
X-ray absorption fine structure (XAFS) above the Fe K-edge and the Ce L3-edge in amorphous Al90Fe(x)Ce10-x (x = 3, 5, and 7) alloys have been measured and analyzed. Quantitative analyses of the Fe K-edge and Ce L3-edge EXAFS spectra are limited to local structure parameters of the first coordination sphere. Using a theoretical multiple scattering (MS) approach, we show that the Fe and Ce XANES are sensitive to the structure of coordination spheres, which extend up to nearly 4.5 A. Comparison of experimental XANES spectra with theoretical MS results allows one to determine the local structure around the iron and cerium sites up to at least the third shell of atoms.
