FUNCTIONAL IMMUNOLIPOSOMES HARBORING A BIOSYNTHETICALLY LIPID-TAGGED SINGLE-CHAIN ANTIBODY

Citation
Ml. Laukkanen et al., FUNCTIONAL IMMUNOLIPOSOMES HARBORING A BIOSYNTHETICALLY LIPID-TAGGED SINGLE-CHAIN ANTIBODY, Biochemistry, 33(38), 1994, pp. 11664-11670
Citations number
54
Categorie Soggetti
Biology
Journal title
ISSN journal
00062960
Volume
33
Issue
38
Year of publication
1994
Pages
11664 - 11670
Database
ISI
SICI code
0006-2960(1994)33:38<11664:FIHABL>2.0.ZU;2-Q
Abstract
An anti-2-phenyloxazolone single-chain antibody was expressed in Esche richia coli as a lipoprotein fusion in order to generate a biosyntheti cally lipid-tagged molecule [Laukkanen et al. (1993) Protein Eng. 6, 4 49-454]. For purification, a hexahistidinyl tag was introduced to the C-terminus of the protein. The resulting antibody, termed Ox lpp-scFv- H6, was membrane-bound, displayed hapten-binding activity, and contain ed the lipoprotein-specific lipid modification as indicated by metabol ic [H-3]palmitic acid labeling. The Ox lpp-scFv-H6 was purified by imm obilized metal affinity chromatography followed by hapten-based affini ty chromatography to essential homogeneity with a yield of 0.4-1.6 mg/ L of culture. In detergent dialysis, the purified antibody partitioned quantitatively into phospholipid liposomes. The immunoliposome prepar ation consisting of a homogeneous population of unilamellar 100-200 nm vesicles displayed specific hapten-binding activity as measured by us ing ELISA and surface plasmon resonance (SPR)-based realtime biospecif ic interaction analysis. In SPR experiments, the immunoliposomes exhib ited virtually irreversible binding to immobilized hapten compared to soluble antibody fragments, consistent with the predicted multivalent binding. Biosynthetic lipid-tagging of antibodies may prove useful for immunoliposome-based diagnostic and therapeutic applications.