In vitro α-complementation of β-galactosidase on a bacteriophage surface

Abstract

Surface display of large multimeric non-secreted proteins is advantageous on the bacteriophage λ compared with the widely used filamentous phage systems. A model system, the α-complementation of β-galactosidase, was used for both further general characterization of protein-protein interactions on the λ tail tube surface and for specifically probing the structure of the phage-displayed β-galactosidase tetramer. In this complementation system, dimeric enzymatically inactive N-terminal deletion mutants of β-galactosidase (α-acceptors) interact with peptides whose sequences span the region of the deletion (α-peptides) with the subsequent formation of tetramers and restoration of activity. The λ phage could tolerate incorporation into their tail tubes of a limited number of copies of V protein (gpV) subunits C-terminally modified with an active α-peptide. Purified α-peptide phage showed specific in vitro α-complementation with an α-acceptor extract; the features of this reaction suggested that each complemented monomer can directly associate with an α-peptide displayed within the same tail tube structure. In contrast to the α-peptide, attempts to surface display an α-acceptor protein in a similar manner were unsuccessful. The implications of this work for surface-display cDNA libraries are discussed.