Department of Biochemistry, University of Washington, Seattle, United States; Institute for Protein Design, University of Washington, Seattle, United States; Eugene McDermott Center for Human Growth and Development, University of Texas Southwestern Medical Center, Dallas, United States; Department of Biophysics, University of Texas Southwestern Medical Center, Dallas, United States
Amy L Schaefer
Department of Microbiology, University of Washington, Seattle, United States
Emily Kenna MacLeod
Department of Microbiology, University of Washington, Seattle, United States
Angelina Zimenko
Department of Microbiology, University of Washington, Seattle, United States
Department of Biochemistry, University of Washington, Seattle, United States; Institute for Protein Design, University of Washington, Seattle, United States; Howard Hughes Medical Institute, University of Washington, Seattle, United States
Many bacteria communicate with kin and coordinate group behaviors through a form of cell-cell signaling called acyl-homoserine lactone (AHL) quorum sensing (QS). In these systems, a signal synthase produces an AHL to which its paired receptor selectively responds. Selectivity is fundamental to cell signaling. Despite its importance, it has been challenging to determine how this selectivity is achieved and how AHL QS systems evolve and diversify. We hypothesized that we could use covariation within the protein sequences of AHL synthases and receptors to identify selectivity residues. We began by identifying about 6000 unique synthase-receptor pairs. We then used the protein sequences of these pairs to identify covariation patterns and mapped the patterns onto the LasI/R system from Pseudomonas aeruginosa PAO1. The covarying residues in both proteins cluster around the ligand-binding sites. We demonstrate that these residues are involved in system selectivity toward the cognate signal and go on to engineer the Las system to both produce and respond to an alternate AHL signal. We have thus demonstrated that covariation methods provide a powerful approach for investigating selectivity in protein-small molecule interactions and have deepened our understanding of how communication systems evolve and diversify.
