Fig. 2: Identification of hypervariable regions in Gp38 adhesin and their receptors.

An alignment of identified Gp38 adhesin proteins in our collection to the reference adhesin protein of Escherichia phage vB_EcoM-fFiEco06, NC_054914.1:156717-157517 was performed in CLC, showing the typical modularity of Gp38 in our phage collection, with a conserved N- and C-terminus (gray and green respectively), which are flanking the alternating conserved GRMs (blue) and diverse HVS (red) (a). The position of the different protein modules is indicated in an AlphaFold prediction of phage FL20, in which the N- and C-terminus (gray and green) are building the attachment domain and beta-helix domain, and the GRM and HVS (blue and red) are building the polyglycine rich sandwich domain, important for binding (b). The variability of gp38 adhesin genes was analyzed by constructing a phylogenetic tree excluding their conserved N-terminus. Receptor recognition of the isolated phages carrying Gp38 adhesin was analyzed by spotting phage dilutions on MG1655 deletion mutants lacking common phage receptors, i.e., ompA, ompF, tsx, tonB, btuB, fadL, lamB, tolC, ompW, and ompC. Numbers display the log of the EOP (efficiency of plaquing) to show infection reduction. No infection or reduced infection (defined as more than three log reduction) is highlighted according to the deleted gene. For FL31, only a one log reduction could be observed for ompF and tolC deletion mutants, which is a higher reduction than for any other deletion mutant but lower than the threshold. Individual Gp38 variants were ordered according to a phylogenetic tree built using the Gp38 sequences lacking the conserved N-terminus (c).