Journal of Lipid Research, Vol. 24, 245-252, March 1983
Copyright © 1983 by Lipid Research, Inc.

Selective proteolytic digestion as a method for the modification of human HDL₃ structure

John B. Swaney

Departments of Biochemistry and Medicine, Albert Einstein College of Medicine, Bronx, NY 10461

Trypsin digestion of human high density lipoprotein (d 1.125-1.21 g/ml) on which the lysine residues have been masked with the reversible blocking group, 2,3,4,5-tetrahydrophthallic anhydride (THPA), was found to result in the fragmentation of the apoA-I component, but not the apoA-II component of this lipoprotein particle. Approximately 50-80% of the apoA-I polypeptide was found in a lipid-free fraction, while the residual apoA-I material plus the apoA-II moiety constituted a core particle that contained most of the original lipid. Immunological analysis indicated that such fragmentation did not affect the immunoreactivity of apoA-II, but that all immunoreactivity of apoA-I was lost within the first 30 min of trypsinization. By column chromatography and electron microscopy this core particle appeared identical in size with the untrypsinized THPA-modified HDL₃ material. Size analysis of the core particle peptides suggests that not all of the A-I molecules present on the HDL₃ are trypsinized to the same extent, which indicates possible nonequivalence of these peptide chains. Analysis of the amino acid composition revealed a somewhat greater proportion of hydrophobic residues in the lipid-bound fraction than in the lipid-free fraction. Analysis of tryptophan showed that almost all of this highly hydrophobic residue was found in the lipid-bound fraction; this suggests that lipid binding occurs preferentially in the more hydrophobic domains of the A-I molecule. Incubation of the core particle with intact apoA-I, obtained from either human or bovine HDL, showed that these proteins could be incorporated to regenerate an HDL₃ of selectively altered protein composition, compared to the original lipoprotein. It is concluded that some latitude is allowable in the surface/volume relationship in lipoproteins before reorganization of the particle is required; this might, for example, provide a mechanism whereby the HDL could serve in a storage role for the C apolipoproteins in plasma.—Swaney, J. B. Selective proteolytic digestion as a method for the modification of human HDL₃ structure.

Supplementary key words trypsinization • apoA-I • apoA-II

Submitted on June 2, 1982
Revised on November 3, 1982

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