Activity-based protein profiling (ABPP) is a chemoproteomic tool for detecting active enzymes in complex biological systems. We used ABPP to identify secreted bacterial and host serine hydrolases that are active in animals infected with the cholera pathogen Vibrio cholerae. Four V. cholerae proteases were consistently active in infected rabbits, and one, VC0157 (renamed IvaP), was also active in human choleric stool. Inactivation of IvaP influenced the activity of other secreted V. cholerae and rabbit enzymes in vivo, and genetic disruption of all four proteases increased the abundance of intelectin, an intestinal lectin, and its binding to V. cholerae in infected rabbits. Intelectin also bound to other enteric bacterial pathogens, suggesting that it may constitute a previously unrecognized mechanism of bacterial surveillance in the intestine that is inhibited by pathogen-secreted proteases. Our work demonstrates the power of activity-based proteomics to reveal host-pathogen enzymatic dialog in an animal model of infection.
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We gratefully acknowledge R. LaRocque, Y. Millet and J. Lee for reagents and technical assistance, and members of M.K.W.'s lab for helpful discussions. We also thank V. Carey for the statistical analysis of MS data and D. Bak for help formatting MS data sets. This work was supported by the National Institutes of Health (R37 AI-042347 to M.K.W., F31 AI-120665 to T.H., R01 AI-106878 and U01 AI-058935 to E.T.R., Institutional Training Grant T32 DK 7477-30 to S.K.H.), the Howard Hughes Medical Institute (M.K.W.), the Charles A. King Trust Postdoctoral Fellowship Program, Bank of America, N.A., Co-Trustee (S.K.H.), the Damon Runyon Cancer Research Foundation (DRR-18-12 to E.W.), the Smith Family Foundation (E.W.) and the Swiss National Science Foundation (P300P3_155287/1 to S.A.).
The authors declare no competing financial interests.
Supplementary Results, Supplementary Tables 1–9 and Supplementary Figures 1–10. (PDF 7161 kb)
FP-biotin-enriched proteins detected in wild-type cecal fluid, comparative ABPP-MudPIT analysis of wild-type and S361A cecal fluid and combined unfiltered data from all ABPP-MudPIT analyses of wild-type cecal fluid. (XLSX 92 kb)
FP-biotin–enriched proteins from human choleric stool supernatant and unfiltered data from ABPP-MudPIT analysis of human choleric stool supernatant. (XLSX 28 kb)
ABPP-MudPIT analysis of wild-type and ΔivaP biofilm culture supernatants. (XLSX 23 kb)
ABPP-MudPIT analysis of exponential- and stationary-phase V. cholerae cell lysates. (XLSX 40 kb)
Comparative total and free fatty acid analysis of wild-type and Δquad cecal fluid. (XLSX 21 kb)
V. cholerae and rabbit proteins detected in unfractionated wild-type and Δquad cecal fluid. (XLSX 188 kb)
V. cholerae and human proteins detected in unfractionated human choleric stool. (XLSX 52 kb)
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Hatzios, S., Abel, S., Martell, J. et al. Chemoproteomic profiling of host and pathogen enzymes active in cholera. Nat Chem Biol 12, 268–274 (2016). https://doi.org/10.1038/nchembio.2025
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