Receptor binding protein of prophage reversibly recognizes the low-molecular weight subunit of the surface-layer protein SlpA in Clostridioides difficile

Abstract

Receptor-binding proteins (RBPs) are located at the viral tail and mediate the initial recognition of phage to a specific bacterial host. Phage RBPs have co-evolved with numerous types of host receptors resulting in the formation of a diverse assortment of cognate pairs of RBP-receptors that function during the phage attachment step. Although several <i>Clostridioides difficile</i> bacteriophages have been discovered, their RBPs are poorly described. Using homology analysis, putative prophage-tail structure (pts) genes were identified from the prophage genome of the <i>C. difficile</i> HN10 strain. Competition and enzyme-linked immunosorbent assays, using recombinant Pts_HN10M, demonstrated the interaction of this Pts to <i>C. difficile</i> cells, suggesting a role as a phage RBP. Gel filtration and cross-linking assay revealed the native form of this protein as a homotrimer. Moreover, truncated variants indicated that the C-terminal domain of Pts_HN10M was important for binding to <i>C. difficile</i> cells. Interaction of Pts_HN10M was also observed to the low-molecular weight subunit of surface-layer protein A (SlpA), located at the outermost surface of <i>C. difficile</i> cells. Altogether, our study highlights the function of Pts_HN10M as an RBP and potentially paves the way toward phage engineering and phage therapy against <i>C. difficile</i> infection.