This disclosure relates to the discovery and isolation of the entire cluster of genes encoding R-type high molecular weight bacteriocins that specifically kill Clostridium difficile bacteria, dangerous pathogens. Also disclosed are methods of producing the R-type bacteriocins in innocuous aerobic producer cells. Disclosed also are small, non-ORF1374 receptor binding domains (RBDs), which are incorporated into diffocins to form engineered or variant diffocins having altered killing spectra. Variant diffocins provided herein may include a heterologous RBD and its cognate base plate attachment region (BPAR), or a fused BPAR. This invention offers a potent bactericidal agent with increased thermal and pH stability, and methods for producing it, in order to kill selectively C. difficile bacteria in the environment of the gastrointestinal tract where they can cause great harm and even death of the infected patient or farm animal.
