Cation-chelation and pH induced controlled switching of the non-fouling properties of bacterial crystalline films.

We report the controlled loss of the anti-fouling activity of the S-layer protein SbpA from Lysinibacillus sphaericus (CCM2177). This protein forms crystal-like films with square lattice (p4) via self-assembly on almost any type of surfaces. Such engineered bioinspired nanometric membranes are known by their excellent preventive performance under biological conditions. However, their exposure to certain treatments can lead to gradual degradation of the S-protein layer. In this work, two distinctive approaches are studied for understanding either specific or non-specific degradation of the film, by treatment with a chelating agent (EDTA), which interacts with inner Ca2+ ions, or Citrate buffer (with pH<pI), respectively. Subsequently, the degraded protein films have been tested upon binding of polyelectrolytes of different charge and endothelial HUVEC cells, and their performance compared to that of intact S-layers. The SbpA protein layer degradation process as well as its impact on the loss of anti-fouling properties have been characterized, in terms of mass and structural changes, by means of real time quartz crystal microbalance with dissipation (QCM-D) monitoring, atomic force microscopy (AFM) experiments, and fluorescence microscopy. The results show that overall structure degradation (citrate buffer) has a higher impact on the loss of antifouling properties than selective removal of divalent cations. Thus, crystal structure integrity is a necessary condition for bacterial antifouling properties.