Reconstitution of purified detergent-soluble HLA-A and HLA-B antigens into phospholipid vesicles.
- 1 July 1978
- journal article
- research article
- Published by Proceedings of the National Academy of Sciences in Proceedings of the National Academy of Sciences
- Vol. 75 (7), 3230-3234
- https://doi.org/10.1073/pnas.75.7.3230
Abstract
Purified detergent-soluble human histocompatibility antigens (HLA-A and HLA-B) were reconstituted into phospholipid vesicles by mixing the protein and lipid together in the presence of octylglucoside (octyl-.beta.-D-glucopyranoside) or deoxycholate and removing the detergent by dialysis. The resulting preparation consisted of lipid vesicles containing all or most of the added protein. The protein in the vesicles was antigenically active, as demonstrated by specific binding to anti-.beta.2-microglobulin Ig[immunoglobulin]G-Sepharose beads and by specific inhibition of alloantibody and complement-mediated cytotoxicity. Protein incorporated into vesicles at a protein/phospholipid ratio of 1:10 showed an asymmetric distribution of the HLA-A and HLA-B molecules, with virtually all of the antigens oriented facing the external medium. Cleavage experiments with proteases showed that the molecule was attached to the vesicle membrane via the COOH terminus, consistent with its proposed structure in intact cellular plasma membranes. Electron micrographs of the vesicles showed 50-60 .ANG. knobs on the outer surface similar to structures observed for other membrane proteins. HLA-A and HLA-B could also be incorporated into vesicles together with Semliki Forest virus membrane proteins. The resulting preparations should be useful in defining the molecular interactions involving HLA-A and HLA-B antigens in the immune response.This publication has 22 references indexed in Scilit:
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