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 6,000 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.
All data generated or analyzed during this study are include in the manuscript and supporting files. Source data files have been provided for the protein sequences analyzed and for Figure 6, Figure 4 - figure supplement 2, Figure 5 - figure supplement 1, and Figure 7 - figure supplement 2.
- E Peter Greenberg
- Qian Cong
- Qian Cong
- Samantha Wellington Miranda
- David Baker
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
- Michael T Laub, Massachusetts Institute of Technology, United States
- Received: April 6, 2021
- Accepted: June 25, 2021
- Accepted Manuscript published: June 28, 2021 (version 1)
© 2021, Wellington Miranda et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.