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    Structural Changes in Hemoglobin during Adsorption to Solid Surfaces: Effects of pH, Ionic Strength and Ligand Binding

    Year: 1998

    Journal: Proc. Natl. Acad. Sci.USA (Biophysics) 95 (1998) 12271-12, 20100827

    Authors: Höök F. 1 2, Rodahl M. 2, Kasemo B. 2, Brzezinski P. 1

    Last authors: Peter Brzezinski

    Organizations: 1 Department of Biochemistry and Biophysics, Göteborg University and Chalmers University of Technology, Medicinaregatan 9C, SE-413 90 Göteborg, Sweden 2 Department of Applied Physics, Chalmers University of Technology, SE-412 96 Göteborg, Sweden

    Country: Sweden

    We have studied the adsorption of two structurally similar forms of hemoglobin (met-Hb and HbCO) to a hydrophobic surface, using a novel Quartz Crystal Microbalance (QCM) technique. This technique allows simultaneous, time-resolved measurements of changes in frequency (f) (c.f. mass) and energy dissipation (D) (c.f. rigidity/viscoelastic properties) of the QCM during the adsorption process, which makes it possible to investigate the viscoelastic properties of the different protein layers. Below the isoelectric points of both met-Hb and HbCO, the D/ graphs displayed two phases with significantly different slopes, which indicates two states of the adsorbed proteins with different viscoelastic properties. The slope of the first phase was smaller than that of the second phase, which indicates that the first phase was associated with binding of a more rigidly attached, presumably denatured, protein layer, whereas the second phase was associated with formation of a second layer of more loosely-bound proteins. This second layer desorbed upon increase of the pH from 5.0 to 6.5, which the first layer did not, or upon reduction of adsorbed met-Hb and subsequent binding of CO. Thus, the results described above indicate that the adsorbed proteins in the second layer were in a native-like state. This information could only be obtained thanks to simultaneous, time-resolved measurements of changes in D and f, demonstrating that the QCM technique provides unique information about the mechanisms of protein adsorption to solid surfaces.