Identification of the receptor-binding regions of pb5 proteins of bacteriophages T5 and BF23.
The receptor-binding protein pb5(T5) of bacteriophage T5, when expressed from the oad gene cloned in pVK88 under the control of the phage T7 promoter/polymerase system, has been shown to bind to its FhuA receptor on the surface of E. coli, where it blocks FhuA for subsequent adsorption of T5 (Mondigler et al., FEMS Microbiol. Lett., 130, 293-300, 1995). In the present study the blocking assay has been applied to analyze the effects of several mutations within oad on the FhuA-binding properties of corresponding pb5 derivatives. Three classes of mutations were tested: (i) oad deletion derivatives, (ii) the oad mutation known to interfere with FhuA-binding of T5 (Heller and Bryniok, J. Virol., 49, 20-25, 1984), and (iii) linker-insertion mutations at a site very close to the oad mutation. Of the corresponding pb5 derivatives only one, a deletion derivative lacking the 153 C-terminal amino acids, was as active in the blocking assay as wild-type pb5(T5). All other derivatives were inactive or almost inactive. Isolation and molecular characterization of phenotypic revertants of T5oad showed that all revertants were true genotypic revertants of the oad mutation. The oad mutation has been identified as a G to T exchange resulting in a substitution of Gly for Trp at position 166 of pb5(T5). DNA sequencing of the hrs gene of bacteriophage BF23 and comparing the deduced amino acid sequence of pb5(BF23) with that of pb5(T5) revealed distinct regions of similarity and nonsimilarity. We propose that the receptor-binding region of pb5(T5) (pb5(BF23)) is formed by the region of nonsimilarity extending from amino acid position 89 (88) to position 305 (283).
Mondigler
M
M
Fakultät für Biologie, Universität Konstanz, Germany.
Holz
T
T
Heller
K J
KJ
D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D003090:Coliphages / Q000737:chemistry / Q000378:metabolism*; D029968:Escherichia coli Proteins; D015967:Gene Expression Regulation, Viral; D005814:Genes, Viral; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D011991:Receptors, Virus / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship; D013604:T-Phages / Q000737:chemistry / Q000378:metabolism*; D014764:Viral Proteins / Q000737:chemistry / Q000378:metabolism*
D001425:Bacterial Outer Membrane Proteins; D029968:Escherichia coli Proteins; C050900:FhuA protein, E coli; D011991:Receptors, Virus; D014764:Viral Proteins; C067129:oad protein, Bacteriophage T5