Functionalized nanoparticles for enhanced affinity precipitation of proteins

A method for purifying a target protein produced by host cells without chromatography, comprising

• (a) separating the target protein from the host cells;
• (b) exposing the separated target protein of step (a) to a nanoparticle in a first solution to form a complex of the target protein and the nanoparticle, wherein the nanoparticle comprises a fusion protein and a scaffolding domain, wherein the fusion protein is covalently bound to the scaffolding domain, wherein the fusion protein comprises an affinity domain capable of binding specifically the target protein and a stimuli responsive precipitation domain, wherein the nanoparticle is bound specifically to the target protein in the first solution, wherein the scaffolding domain comprises self-assembled proteins and has a diameter of at least 10 nm, wherein the stimuli responsive precipitation domain is an elastin-like polypeptide (ELP) and the scaffolding domain is an E2 core of the pyruvate dehydrogenase complex (E2), wherein the first solution has a salt concentration of 50-200 mM and a pH of 6-9, whereby the complex precipitates out of the first solution;
• (c) adding the precipitated complex of step (b) to a second solution having a pH less than 4, whereby the precipitated complex of step (b) is solubilized and the target protein is released from the nanoparticles in the second solution;
• and
• (d) applying a stimulus to the second solution of step (c), wherein the stimulus comprises increasing the salt concentration of the second solution, whereby the solubilized nanoparticle of step (c) precipitates out of the second solution and the released target protein of step (c) is purified from the host cells.