Efficient expression and immunoaffinity purification of human trace amine-associated receptor 5 from E. coli cell-free system

G protein-coupled receptors (gpcrs) represent attractive targets for bioactive and drug discovery programs. the availability of purified receptors in milligram quantities is essential to spur the advancement of protein-based analyses in these programs,although it is still a challenging goal to achieve. here we report the production of a bioengineered gpcr of human trace amine-associated receptor 5 (htaar5) from an e. coli cell-free system. both the htaar5 and htaar5-t4 lysozyme fusion proteins (htaar5-t4l) were cloned and expressed in this process,with the latter designed for further protein crystallization trials. the detergent brij-35 was found to solubilize the produced htaar5 and htaar5-t4l effectively. immunoaffinity purification in combination with gel filtration was employed to purify the receptors to high homogeneity. the final yields of monomeric htaar5 and htaar5-t4l from a 1 ml cell-free reaction were 0.4 mg and 0.5 mg,respectively. circular dichroism (cd) spectroscopy indicated that both htaar5 and htaar5- t4l were correctly folded after purification,with characteristic high -helical contents (>45%). © 2013 bentham science publishers.