One-step metal affinity purification of histidine-tagged proteins by temperaturetriggered precipitation. Biotechnol Bioeng 82:605–611

Abstract: The feature of elastin-like proteins (ELPs) to reversibly precipitate above their transition temperature was exploited as a general method for the purification of histidine (His)-tagged proteins. The principle of the single-step metal-affinity method is based on coordi-nated ligand-bridging between the modified ELPs and the target proteins. ELPs with repeating sequences of [(VPGVG)2(VPGKG)(VPGVG)2]21 were synthesized and the free amino groups on the lysine residues were modi-fied by reacting with imidazole-2-carboxyaldehyde to in-corporate the metal-binding ligands into the ELP bio-polymers. Biopolymers charged with Ni2+ were able to interact with a His tag on the target proteins based on metal coordination chemistry. Purifications of two His-tagged enzymes, -D-galactosidase and chlorampheni-col acetyltransferase, were used to demonstrate the util-ity of this general method and over 85 % recovery was observed in both cases. The bound enzymes were easily released by addition of either EDTA or imidazole. The recovered ELPs were reused four times with no observ-able decrease in the purification performance. © 200