Mapping the Functional Landscape of the Receptor Binding Domain of T7 Bacteriophage by Deep Mutational Scanning

Abstract

The interaction between a bacteriophage and its host is mediated by the phage’s receptor binding protein (RBP). Despite its fundamental role in governing phage activity and host range, molecular rules of RBP function remain a mystery. Here, we systematically dissect the functional role of every residue in the tip domain of T7 phage RBP using a high-throughput, locus-specific, phage engineering method. We find that function-enhancing mutations are concentrated around outward-facing loops suggesting directionality and orientational-bias in receptor engagement. These mutations are host-specific, indicating adaptation to individual hosts and highlighting a tradeoff between activity and host range. We discover gain-of-function variants effective against resistant strains and host-constricting variants that selectively eliminate certain hosts. We demonstrate therapeutic utility against uropathogenic E. coli by engineering a highly active T7 to avert emergence of spontaneous resistance of the pathogen. Our approach is generalizable to other phages and will enable the design of programmable phages.

ORACLE reveals a bright future to fight bacteria

“…Yet engineering phages to target harmful bacterial species requires scalable genetic tools that can precisely alter the genomes of these viruses. Now, in eLife, Srivatsan Raman and colleagues at University of Wisconsin-Madison – including Phil Huss as first author – report a new method that can create thousands of mutations in a host-specifying phage protein, showing how these changes alter which bacteria the virus can target ( Huss et al, 2021 ).…”