LIPID-TAGGED ANTIBODIES - BACTERIAL EXPRESSION AND CHARACTERIZATION OF A LIPOPROTEIN SINGLE-CHAIN ANTIBODY FUSION PROTEIN

In order to achieve a stable and functional immobilization of antibodi es, we investigated the possibility of adding hydrophobic membrane anc hors to antibody fragments expressed in Escherichia coli. The DNA sequ ence encoding the signal peptide and the nine N-terminal amino acid re sidues of the major lipoprotein of E.coli was fused to the sequence of an anti-2-phenyloxazolone single-chain Fv antibody fragment [Takkinen et al. (1991) Protein Engng, 4, 837-841]. The expression of the fusio n construct in E.coli resulted in specific accumulation of an immunore active 28 kDa polypeptide. Unlike the unmodified single-chain Fv fragm ent, the fusion protein was cell-associated, labelled by [H-3]palmitat e which is indicative of the presence of N-terminal lipid modification , partitioned into the detergent phase upon Triton X-114 phase separat ion and was localized predominantly in the bacterial outer membrane. T he fusion antibody displayed specific 2-phenyloxazolone-binding activi ty in the membrane-bound form and after solubilization with non-ionic detergents. Furthermore, upon removal of detergent the fusion antibody was incorporated into proteoliposomes which displayed specific hapten -binding activity. Our results show that antibodies can be converted t o membrane-bound proteins with retention of antigen-binding properties by introduction of lipid anchors during biosynthesis. This approach m ay prove useful in the design of immunoliposomes and immunosensors.