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Receptor specificity of the Escherichia coli T-even type phage Ox2. Mutational alterations in host range mutants
Abstract:
The T-even type Escherichia coli phage Ox2 uses the outer membrane protein OmpA as a receptor. The protein is recognized with the ends of the virion's long tail fibers. The 266 residue protein 38 is located at this site and acts as an adhesin. Host-range mutants had previously been isolated from Ox2. Mutant Ox2h5 is able to infect cells possessing an altered OmpA protein, which renders the cell resistant to Ox2. Ox2h10 was selected from Ox2h5. This phage recognizes the OmpC protein in addition to the OmpA protein. Ox2h12, which stems from Ox2h10, binds to OmpC with high affinity, but has lost efficient binding to OmpA. The mutational alterations caused in genes 38 are: Asp231----Asn(h5) and His170----Arg(h10). The triple mutant Ox2h12 possesses an insertion of a Gly residue next to Gly121. The three mutants have additionally acquired mutations affecting their base plate, making them "trigger-happy". When protein 38 was compared with the same protein derived from other E. coli phages, it was found to contain two constant and one variable domains, the latter harboring four hypervariable regions flanked by a largely conserved glycine-rich sequence. The h5 and h10 mutations occurred within two hypervariable areas, while the additional Gly residue was present in one of the flanking conserved sequences. On the basis of these results, as well as those obtained from host-range mutants analyzed previously, a model for such adhesins is proposed. Receptor recognition is most likely performed via the hypervariable regions, which may form loops held together in close proximity by the oligoglycine sequences. The latter may achieve this by being part of highly compact omega loops.
Insights
Bacteriophage Ox2 adhesin protein 38 mutations alter host range by changing interactions with Escherichia coli outer membrane proteins OmpA and OmpC. These changes provide insights into phage-adhesin evolution and receptor binding mechanisms.
Area of Science:
- Microbiology
- Molecular Biology
- Structural Biology
Background:
- T-even Escherichia coli phage Ox2 utilizes outer membrane protein A (OmpA) as its receptor, with the adhesin protein 38 mediating this interaction.
- Host-range mutants of Ox2 have been previously isolated, indicating adaptability in receptor binding.
Purpose of the Study:
- To investigate the molecular basis of altered host-range specificity in specific T-even phage Ox2 mutants.
- To elucidate the role of adhesin protein 38 domains in receptor recognition and binding affinity.
Main Methods:
- Isolation and characterization of host-range mutants Ox2h5, Ox2h10, and Ox2h12.
- Determination of mutational alterations in adhesin protein 38 genes and base plate components.
- Comparative analysis of protein 38 from different E. coli phages to identify conserved and variable domains.
Main Results:
- Mutations Asp231Asn (h5) and His170Arg (h10) in protein 38 conferred altered OmpA binding and enabled recognition of OmpC.
- Mutant Ox2h12 exhibited high-affinity OmpC binding and reduced OmpA binding, with an insertion in a conserved glycine-rich sequence.
- Mutations affected the base plate, leading to increased sensitivity ('trigger-happy') in receptor binding.
Conclusions:
- The variable domains of adhesin protein 38, particularly hypervariable regions, are critical for receptor recognition.
- Oligoglycine sequences likely stabilize these variable regions, forming omega loops that mediate binding to outer membrane proteins like OmpA and OmpC.
- The study proposes a model for adhesin function, highlighting the interplay between hypervariable regions and conserved structural elements in determining phage host range.