Apolipoprotein E3 (ApoE3) is an important
apolipoprotein in plasma and plays a critical role in
lipid transport and
cholesterol homeostasis. As the only
natural source of this protein,
human blood cannot provide large-scale ApoE3 for research and applications. Therefore, in our study, a
Pichia pastoris expression system was first used to obtain a high-level expression of
secreted, recombinant human ApoE3 (rhApoE3). The full-length sequence encoding ApoE3, gained by
RT-PCR, was inserted into the pPICZalphaC vector and transformed into P. pastoris strain X33, and then the high expression transformants with zeocin resistance were obtained. The growth conditions of the transformant strains were optimized in 50ml conical tubes including pH and inducing time. After induction with
methanol, the expression level of rhApoE3 was 120mg/L in 80L fermentor. RhApoE3 was purified more than 94% purity using SP
Sepharose ion exchange chromatography and source 30RPC. A preliminary biochemical characterization of purified rhApoE3 was performed by analyzing the ability of inhibiting PDGF-induced proliferation of rat
coronary artery smooth muscle cells (SMCs), and the results demonstrated that the function of purified rhApoE3 was similar to natural human ApoE3.
