Conformational Analysis of Serum Apolipoprotein AII in Lipoprotein Complexes with Bifunctional Crosslinking Reagents

Abstract

Apolipoprotein AII isolated from human serum high density lipoproteins was recombined with [soybean] phosphatidylcholine to yield homogeneous particles of 80-120 .ANG. diameter. The radioactive bifunctional crosslinkers dimethyl [1,1''-14C]suberimidate and dimethyl 4,4''-dithio-bis([1-14C]butyrimidate) were reacted with these particles. The kinetics of the reactions and the localization of the crosslinked Lys of the polypeptide chains were determined. Thermolysin [EC 3.4.24.4] hydrolysis followed by 2-dimensional separation of the peptides and isolation of the mono- and bifunctionally modified peptides allowed the assignment of the crosslinked peptides of the apoprotein AII sequence. The crosslinking pattern indicates a close-neighbor relationship (13-15 .ANG.) of the peptide chains between amino acid residues 3, 23, 46 and 55 of the symmetrical halves of the apo AII molecule. A reconstruction of the secondary structure of Apo AII in the lipoprotein complex on the basis of theoretical calculations is given and correlated with the chemical data.