In vivo kinetics as a sensitive method for testing physiologically intact human recombinant apolipoprotein A-I: comparison of three different expression systems.

In order to assess the structural and functional integrity of recombinant human apoA-I, we expressed apoA-I using three different expression systems: Baculovirus transfected Spodoptera frugiperda (Sf9) cells, stably transfected Chinese hamster ovary (CHO) cells, and transformed Escherichia coli (E. coli). Purified apoA-I from the three expression systems was radioiodinated and their catabolism was compared in normolipemic rabbits. The kinetic turnover studies of radiolabelled apoA-I in normolipemic rabbits revealed that highly purified recombinant apoA-I had an identical decay curve compared to native apoA-I, regardless whether it was purified from Sf9 cells, CHO cells, or E. coli. We also determined the association of the three recombinant apoA-I forms with both rabbit and human HDL. All three recombinant apoA-I forms were associated with HDL2 and HDL3 after injection into the rabbits and after incubation with human serum using both a Superose 6 column separation system and density gradient ultracentrifugation. The addition of the pro-segment or the addition of methionine at the amino-terminal end of apoA-I did not alter its metabolism and association to HDL. In conclusion, all studied expression systems are capable of producing high levels of physiologically intact recombinant human apoA-I. The aminoterminal addition of the prosegment of apoA-I or methionine did not alter the in vivo metabolism of apoA-I or its association to HDL.