Delineating the site of interaction on the pIII protein of filamentous bacteriophage fd with the F-pilus of Escherichia coli.
The minor coat protein pIII at one end of the filamentous bacteriophage fd, mediates the infection of Escherichia coli cells displaying an F-pilus. pIII has three domains (D1, D2 and D3), terminating with a short hydrophobic segment at the C-terminal end. Domain D2 binds to the tip of F-pilus, which is followed by retraction of the pilus and penetration of the E. coli cell membrane, the latter involving an interaction between domain D1 and the TolA protein in the membrane. Surface residues on the D2 domain of pIII were replaced systematically with alanine. Mutant virions were screened for D2-pilus interaction in vivo by measuring the release of infectious virions from E. coli F(+) cells infected with the mutants. A competitive ELISA was developed to measure in vitro the ability of mutant phages to bind to purified pili. This allowed the identification of amino acid residues involved in binding to F and to EDP208 pili. These residues were found to cluster on the outer rim of the 3D structure of the D2 domain, unexpectedly identifying this as the F-pilus binding region on the pIII protein.
Deng
Lih-Wen
LW
Cambridge Centre for Molecular Recognition, Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB2 1GA, UK.
Perham
Richard N
RN
D017104:Bacteriophage M13 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D001667:Binding, Competitive; D019921:Biotinylation; D036022:Capsid Proteins; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000378:metabolism*; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed / Q000235:genetics; D009154:Mutation / Q000235:genetics; D018933:Pili, Sex / Q000378:metabolism*; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D020860:Subtilisin / Q000378:metabolism; D014760:Viral Fusion Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D014771:Virion / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000502:physiology
D036022:Capsid Proteins; D004268:DNA-Binding Proteins; D014760:Viral Fusion Proteins; D020860:Subtilisin