Superheated droplets for protein thermal stability analyses of GFP, BSA and Taq-polymerase

Abstract

Here we describe a novel method for the study of protein thermal stability using superheated aqueous samples within virtual reaction chambers. Virtual reaction chambers consist of an aqueous sample droplet encapsulated by an oil droplet on a hydrophobic surface. Such samples can be superheated due to the lack of nucleation sites. The thermal denaturation of proteins is induced through the application of a temperature gradient using a bespoke silicon heating chip. The unfolding of proteins is followed through the addition of a hydrophobic dye that attaches to protein hydrophobic domains that become exposed during denaturation. Using this method, we investigated the thermal stability of green fluorescence protein and Taq-polymerase. A possible screening application of the method was demonstrated by evaluating the effect of ionic concentration on the thermal stability of bovine serum albumin.