Lipid-Protein Interactions between Human Apolipoprotein A-I and Defined Sphingomyelin Species. A13C-NMR Spectroscopic Study

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Abstract
Chromatographically and immunologically homogeneous apolipoprotein A-I (apoLp A-I) from human serum was recombined in separate experiments with 3 species of sphingomyelin differing in the degree of saturation: stearoyl (18:0), oleoyl (18:1) and linoleoyl (18:2). The lipoprotein complexes formed were purified by CsCl density gradient centrifugation between 1.07-1.09 g/cm3 and by gel filtration. Stearoylsphingomyelin does not recombine with the apoprotein A-I below its phase transition temperature (tc = 41.5.degree. C). The lipoproteins eluted with the following apparent MW: 18:0-sphingomyelin apoLp A-I, 8.0 .times. 105; 18:1-sphingomyelin apoLp A-I, 4.0 .times. 105; and 18:2-sphingomyelin apoLp A-I, 4.0 .times. 105. In EM the particles appear as discs of 160-170 .ANG. diameter and 50-60 .ANG. thickness. Their tendency to form stacked aggregates of discs decreases with the degree of their unsaturation. Circular dichroism measurements underline the considerable increase in .alpha.-helicity of the secondary structure of apo A-I after recombination with the phospholipids. This increase in order is equal for the 3 sphingomyelin species (.alpha.-helicity of apoLP A-I = 0.46, after recombination 0.89). In the 3 sphingomyelin species are used in equal molar amounts in the recombination experiment, no preference for any 1 sphingomyelin species is observed. Recombination of apoLp A-I with sphingomyelin, labeled with 13C in the choline group, C-14 of stearic or linoleic or C-11 of oleic acid, were performed for spin lattice relaxation time (T1) experiments. Compared with sphingomyelin liposomes, the polar head groups of these lipids in the lipoprotein particles possess a considerably higher mobility, whereas the changes in T1-times of the C-atoms in the center of the fatty acid chains of the lipids refer to their interactions with the polypeptide side chains. A model of the lipoprotein complexes formed is proposed on the basis of the experimental data.