Polyethyleneimine-mediated chemical extraction of cytoplasmic his-tagged inclusion body proteins from Escherichia coli

Abstract

The selectivity of polyethyleneimine (PEI) in DNA precipitation during chemical extraction was investigated. Chemical extraction was used to recover two His-tagged model proteins: gloshedobin, a thrombin-like enzyme from snake venom, and IbpA, a molecular chaperone, which were expressed mainly in the form of inclusion bodies. High DNA removal efficiency (more than 90%) was achieved at various cell densities (with OD600 ranging from 30 to 150) without affecting the solubility of host cell proteins. Compared to spermine-induced precipitation method reported elsewhere, PEI provided a higher DNA precipitation efficiency at a significantly lower cost. Moreover, PEI obviated the use of EDTA, which has been reported to be essential for the chemical extraction methods, hence exhibiting dual roles in replacing cost-prohibitive spermine and EDTA. The residual PEI in the post-extraction mixture was efficiently counteracted by addition of Mg2+, allowing the streamlined application of the extraction broth to immobilized metal affinity chromatography. Taken together, the PEI-mediated chemical extraction method provides a simpler and more economically viable processing route for the production of recombinant proteins whose expression is hampered by IB formation. © 2008 American Chemical Society and American Institute of Chemical Engineers.