Possibilities of X-Ray Absorption Spectroscopy in the Total External Reflection Geometry for Studying Protein Films on Liquids

XANES spectra of protein films (hemoglobin and alkaline phosphatase) formed on the surface of a liquid subphase in a Langmuir trough have been obtained experimentally. The potential of X-ray absorption spectroscopy in the total external reflection geometry is demonstrated by the example of a protein film based on hemoglobin subjected to the action of urea. It is established that the presence of 0.09 M urea solution in the subphase enhances significantly the ability of hemoglobin to bind zinc and iron ions. Information about the local atomic environment of zinc ions bound with a hemoglobin molecule is obtained. It is shown that each zinc ion is coordinated by four ligands, two of which are amino acid residues of cysteine and histidine. The general concepts of the molecular mechanisms of accumulation of metal ions under the action of disturbing factors are formulated.