Cloning, Expression, Purification And Characterization Of Human Apolipoprotein A-v

As an independent risk factor in the development of coronary arteriosclerosis, elevated level of triglycerides has close relationship with coronary heart disease, myocardial infraction, diabetes and other common diseases. ApoAV, found in the member of Apoa I /c III/a IV gene cluster, was highly conserved in human, dog, rabbit, mouse and birds. Secreted sorely by liver, ApoAV has low concentration in plasma (about 0.1% as compared to ApoA1) but plays an important role in reducing the levels of plasma triglyceride. It is suggested that ApoAV accelerated hydrolysis of plasma triglyceride-rich lipoproteins by interacting with proteoglycan-bound lipoprotein lipase.It is hard to obtain large amount of soluble ApoAV protein directly from animals, therefore few biochemical studies has been carried out in vitro and most experiments were focused on gene analysis. We are aimed to express soluble ApoAV in Escherichia coli by trnasforing two fusion exression plasmids pSJ5-ApoAV and ppSUMO-ApoAV into BL21(DE3) or Rosetta(DE3). After optimizing the culture temperature, induced time, and the concentration of IPTG, we were able to express ApoAV in soluble form at very high level. The recombinant ApoAV proteins from two different plasmids were separated by Ni2+ affinity column, protease digestion and anion exchange column. The CD analysis shows that the purified ApoAV protein has significantα-helix andβ-sheet, which suggested that it may have anα+βstructure.