Role of the C-terminal Modules of Klebsiella Phage KP32 Receptor-Binding Protein gp38 in Protein and Phage Functionality

Despite significant progress in understanding phage biology and their clinical applications, the specificity of phages remains only merely understood and a matter of empirical testing. This study investigates the receptor-binding protein KP32gp38 of Klebsiella phage KP32, which features a unique C-terminal tandem of a carbohydrate-binding module (CBM) and a lectin-like (LEC) domain. We dissected the roles of these modules in trimerization, substrate binding, and specificity. Our results demonstrate that the LEC domain is essential for trimerization, while the CBM domain is crucial for enzymatic activity and capsule binding. Engineered phages lacking these domains confirmed the necessity of both CBM and LEC for full functionality. This work underscores the versatility and evolutionary adaptation of CBM and LEC folds in phage RBPs, providing valuable insights into phage specificity mechanisms. Our findings offer a blueprint for understanding the molecular determinants of phage-host interactions, crucial for advancing phage-based antibacterial therapies.