Refolding of lactate dehydrogenase by zeolite beta

Abstract

We used zeolite beta as an adsorbing matrix to refold recombinant lactate dehydrogenase (LDH) protein collected as an insoluble aggregate from a bacterial expression system. The adsorption isotherm revealed that 1 g ofzeolite adsorbed 200 mg ofdenatured LDH solubi- lized with a buffer containing 6 M of guanidine hydrochloride. The pH of the buffer had little effect on the adsorption, but this property was abolished by preincubation ofthe zeolite with polyethylene glycol (PEG) in a weight ratio of 1:10. These data suggest that the adsorption of LDH depends on the hydrophobicity of the zeolite surface, and that the adsorption of PEG to zeolite is sufficient to release LDH from its surface. LDH was thus released by refolding buffer containing PEG and arginine, and soluble LDH was obtained in its active enzymatic form. The addition ofarginine dramatically increased the yield ofLDH in a dose-dependent manner. The overall refolding efficiency was optimized to 35%. © 2009 American Institute of Chemical Engineers.