IN-VIVO KINETICS AS A SENSITIVE METHOD FOR TESTING PHYSIOLOGICALLY INTACT HUMAN RECOMBINANT APOLIPOPROTEIN-A-I - COMPARISON OF 3 DIFFERENT EXPRESSION SYSTEMS

Citation
Hhj. Schmidt et al., IN-VIVO KINETICS AS A SENSITIVE METHOD FOR TESTING PHYSIOLOGICALLY INTACT HUMAN RECOMBINANT APOLIPOPROTEIN-A-I - COMPARISON OF 3 DIFFERENT EXPRESSION SYSTEMS, Clinica chimica acta, 268(1-2), 1997, pp. 41-60
Citations number
58
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Medical Laboratory Technology",Biology
Journal title
ISSN journal
0009-8981
Volume
268
Issue
1-2
Year of publication
1997
Pages
41 - 60
Database
ISI
SICI code
0009-8981(1997)268:1-2<41:IKAASM>2.0.ZU;2-J
Abstract
In order to assess the structural and functional integrity of recombin ant human apoA-I, we expressed apoA-I using three different expression systems: Baculovirus transfected Spodoptera frugiperda (Sf9) cells, s tably transfected Chinese hamster ovary (CHO) cells, and transformed E scherichia coli (E. coli). Purified apoA-I from the three expression s ystems was radioiodinated and their catabolism was compared in normoli pemic 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 wh ether it was purified from Sf9 cells, CHO cells, or E. coli. We also d etermined the association of the three recombinant apoA-I forms with b oth rabbit and human HDL. All three recombinant apoA-I forms were asso ciated with HDL2 and HDL3 after injection into the rabbits and after i ncubation with human serum using both a Superose 6 column separation s ystem and density gradient ultracentrifugation. The addition of the pr o-segment or the addition of methionine at the amino-terminal end of a poA-I did not alter its metabolism and association to HDL. In conclusi on, all studied expression systems are capable of producing high level s of physiologically intact recombinant human apoA-I. The aminotermina l addition of the prosegment of apoA-I or methionine did not alter the in vivo metabolism of apoA-I or its association to HDL. (C) 1997 Else vier Science BN.