Apolipoprotein (apo) E plays an important role in lipid metabolism, and the 
major isoforms of apoE (apoE2, apoE3, and apoE4) have significantly different 
metabolic effects. Apolipoprotein E4 is associated with a higher risk of both 
heart disease and Alzheimer's disease (AD). Patients homozygous for 
apolipoprotein E2 are predisposed to type III hyperlipoproteinemia, and apoE2 
may be protective against AD. Structure/function studies have proved to be a 
useful tool in understanding how the different apoE isoforms result in different 
pathological consequences. As these studies continue, it is essential to have a 
reliable method to produce large quantities of apoE and mutants of apoE. We 
describe here a method of apoE production in Escherichia coli strain BL21(DE3). 
The cDNA from apoE isoforms was inserted into a pET32a vector with a T7 promoter 
and a fusion partner (thioredoxin). The T7 promoter results in high expression 
of an easily purified His-tagged fusion protein. A thrombin recognition site was 
positioned in the expression vector so that only two novel amino acids (Gly-Ser) 
are added to the amino terminus of apoE following the removal of thioredoxin. 
Approximately 20 mg of apoE is obtained from a 1-liter culture. The major 
isoforms of apoE produced with this system were extensively characterized for 
their ability to bind the low-density lipoprotein (LDL) receptor, for their 
characteristic lipid association preferences, and for their stability as 
measured by guanidine denaturation. The recombinant proteins behaved identically 
to plasma-derived apoE isoforms.