Thermodynamically Reversible Addressing of a Stimuli Responsive Fusion Protein onto a Patterned Surface Template†

The sensitivity of an elastin-like polypeptide (ELP) to environmental stimuli is used to reversibly immobilize a fusion partner, thioredoxin (TRX), onto a hydrophobic surface. An ELP, fused to TRX at its C-terminus, adsorbs onto a hydrophobic self-assembled monolayer (SAM) on gold above its inverse phase transition temperature (Tc) and is resolubilized from the surface when the solution temperature is lowered below Tc. We show that the adsorbed fusion partner TRX is recognized by an antibody specific to TRX and that the complex is also resolubilized from the surface below Tc. Adsorption of TRX-ELP is inhibited by hydrophilic surfaces that are defect-free. In situ ellipsometry shows that the ELP-anchored TRX adsorbs above Tc up to a well-defined layer thickness that is greater than a monolayer and that the adsorption and desorption cycle can be repeated. These results are confirmed by atomic force microscopy and by surface plasmon resonance spectroscopy. The preferential adsorption of TRX-ELP on hydrophobic surfaces is used to create a pattern of adsorbed protein on a surface composed of a pattern of hydrophobic and hydrophilic SAMs on gold. We term this method to reversibly address an ELP fusion protein to chemically distinct regions of a patterned surface by an external stimulus “thermodynamically reversible addressing of proteins” (TRAP).