Protein-surface interactions determine the ultimate fate of biomaterials and the success of biomedical and diagnostic devices. For instance, with simple implants (stents, shunts, and objects with organ-level function such as artificial hearts, and orthopedic implants), the goal is a device surface that is invisible to the body, except at the functioning junctions, where living tissue connects to the device. In practice, such implants can initiate immuno-reactions, blood clots (thrombosis), or provide a site for infection. The extent to which these problems occur depends largely on protein adhesion to implant surfaces, a process which occurs before these surfaces are encountered by cells. More sophisticated biomaterials, such as tissue scaffolds, should direct the growth of new cells towards a desired outcome. Here it would be useful for materials to somehow signal cells and restrict biological response; however, the design of such materials must incorporate the fact that protein-surface interactions determine the ultimate cellular response.
The first proteins to reach a surface adsorb and often denature, and their initial behavior governs the subsequent adsorption of other proteins, cells, and physiological cascades. In the Santore lab, we employ optical and spectroscopic techniques to track protein adsorption at solid-liquid interfaces in carefully-controlled flow conditions. The goal of these fundamental studies is an understanding of factors affecting proteins on surfaces and the mechanisms for interfacial protein behavior, ultimately to facilitate protein manipulation strategies.
Adsorption kinetics. Adsorption kinetics are monitored via fluorescence and optical methods with the time resolution necessary to identify the adsorption mechanism. Adsorption is usually transport-limited, at least at short times, as shown in Figure 2. The initial adhesion of proteins to most surfaces is fast and independent of surface chemistry because the surfaces of proteins contain many different chemical groups, at least a few of which are likely to be attracted to the particular surface being studied.
• C.F. Wertz, and M. Santore, Langmuir 15, 8884 - 8894 (1999).
Surface relaxations and denaturing. Proteins encountering surfaces may reconfigure, from side-on to end-on, or from face up to face down, or they may simply denature on the surface. In the Santore lab, we distinguish between protein reconfiguration and denaturing, and measure the rates of the two processes, as a function of surface chemistry. For instance, we have determined that lysozyme, a relatively small, hard protein “rolls-over” shortly after adsorbing to surfaces, without substantial denaturing. Conversely, fibrinogen, an important blood-clotting protein, denatures with a surface chemistry-dependent rate.
• C. Wertz and M. Santore, Langmuir 18, 1190-1199 (2002).
Biophysics. A careful analysis of fibrinogen denaturing kinetics revealed single exponential relaxation behavior on hydrophobic surfaces. This result comes as a surprise if one likens the interior protein surfaces participating in intramolecular associations with the exterior protein surfaces of ligand-receptor interactions. The latter display complex energy landscapes, a series of kinetic energy barriers, each of which dominates intermolecular adhesion at different adhesion rates. In the case of fibrinogen denaturing, we see evidence for a dominant intra-molecular energy barrier at the rates inherent to un-forced interfacial protein denaturing.
• C. Wertz and M. Santore*, Langmuir, 18, 706-715 (2002).• C. Wertz and M. Santore*, in preparation
Ongoing protein work in the Santore lab includes studies focusing on the biophysics of surface-induced protein unfolding (for instance, using AFM), complex fluid (such as blood) –surface interactions, and separately, on materials designed for protein manipulation. The latter include surfaces with monodisperse nanometer-scale (3-50 nm) patterns of controlled chemistry.One method to produce such nano-patterned surfaces employs phase separated polymer films as templates, in Figure 4.