As can be seen in Fig. 6, the adsorption rate slightly increases at about half the maximum interfacial concentration. Such an increase could be the signature of a structural change at the interface. It was recognized for many years that protein crowding at the interface leads to profound changes there because of the increasing probability of the protein-protein interactions (Andrade 1985). In the present case, such a change in the adsorption rate could also be caused by the conditions of flow in the channel, associated with the early saturation of the surface upward of the point of examination. Simulations suggest, however, that this hypothesis should be rejected, as it would require an adsorption constant much higher than that deduced from the previous analysis (Vasina and Dejardin 2004). To describe the conformational changes (see Chap. 6) and/or orientation, many models can be proposed, among them the side-on/end-on process, the reverse process, and the spreading of the (soft) protein on the surface. Techniques like neutron reflectivity (Su et al. 1998), ellipsometry (Bae et al. 2005; Cuypers et al. 1978; McClellan and Franses 2005; Poksinski and Arwin 2004; Seitz et al. 2005; Werner et al. 1999), and scanning angle reflectometry (Ladam et al. 2002; Schaaf and Déjardin 1988) are able to provide information about the orientation of an ellipsoid-modeled molecule. Circular dichroism and differential scanning calorimetry are also useful in the study of the protein conformations (Giacomelli and Norde 2001; Norde and Zoungrana 1998; Vermeer and Norde 2000; Vermeer et al. 2001; Voegel et al. 1987; Zoungrana et al. 1997). Infrared attenuated total reflection (Noinville et al. 2002a,b, 2003) provides data about possible changes in the tertiary structure (see Chap. 6).
The TIRF technique was refined to take into account the dependence on the emission properties of the fluorophore with its ionization state or with the electric potential (Daly et al. 2003; Robeson and Tilton 1996). If the ionic strength is chosen in such a way that the interfacial electric potential varies over a distance comparable to the protein size, then the signal will be dependent upon the orientation of the molecule. Indeed the interfacial transition is easily observed and is concomitant with the change in kinetic regime.
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