External surface display of proteins linked to DNA-binding domains.
A novel system (DBDX) was developed which allows the external surface display on filamentous bacteriophage of proteins fused to either the N- or the C-terminus of a DNA-binding protein. In conjunction with helper phage infection, expression of proteins fused to the estrogen receptor DNA-binding domain (DBD) in a phagemid vector containing the DNA sequence recognized by the DBD resulted in the production of phage particles which display the fusion protein through the phage pVIII coat on the external surface of the particle. The viability of the technique was established with several model systems: particles displaying the C-terminal domain of N-cadherin or the biotinylation domain of propionyl coenzyme A carboxylase fused to the C-terminus of the DBD were found to be bound specifically by antibody or streptavidin, respectively. Human kappa constant region cDNA was selected from a N-terminal DBD fusion lymphocyte cDNA library after two rounds of selection with anti-kappa antibody. This display system may complement currently available bacterial selection techniques.
McGregor
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Skeletal Metabolism Research Unit, Rowett Research Institute, Greenburn Road, Bucksburn, Aberdeen, AB21 9SB, Scotland.
Robins
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D001435:Bacteriophages / Q000235:genetics / Q000502:physiology*; D001483:Base Sequence; D015820:Cadherins / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D019735:Carbon-Carbon Ligases / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D005822:Genetic Vectors; D006801:Humans; D008969:Molecular Sequence Data; D019151:Peptide Library; D015202:Protein Engineering; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000032:analysis*; D011960:Receptors, Estrogen / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000032:analysis
D015820:Cadherins; D004268:DNA-Binding Proteins; D019151:Peptide Library; D011506:Proteins; D011960:Receptors, Estrogen; D011993:Recombinant Fusion Proteins; D019735:Carbon-Carbon Ligases; C022458:propionyl CoA carboxylase (ATP-hydrolyzing)