NTR 2.0: a rationally engineered prodrug-converting enzyme with substantially enhanced efficacy for targeted cell ablation

Abigail V. Sharrock, Timothy S. Mulligan, Kelsi R. Hall, Elsie M. Williams, David T. White, Liyun Zhang, Kevin Emmerich, Frazer Matthews, Saumya Nimmagadda, Selena Washington, Katherine D. Le, Danielle Meir-Levi, Olivia L. Cox, Meera T. Saxena, Anne L. Calof, Martha E. Lopez-Burks, Arthur D. Lander, Ding Ding, Hongkai Ji, David F. AckerleyJeff S. Mumm

Research output: Contribution to journalArticlepeer-review

Abstract

Transgenic expression of bacterial nitroreductase (NTR) enzymes sensitizes eukaryotic cells to prodrugs such as metronidazole (MTZ), enabling selective cell-ablation paradigms that have expanded studies of cell function and regeneration in vertebrates. However, first-generation NTRs required confoundingly toxic prodrug treatments to achieve effective cell ablation, and some cell types have proven resistant. Here we used rational engineering and cross-species screening to develop an NTR variant, NTR 2.0, which exhibits ~100-fold improvement in MTZ-mediated cell-specific ablation efficacy, eliminating the need for near-toxic prodrug treatment regimens. NTR 2.0 therefore enables sustained cell-loss paradigms and ablation of previously resistant cell types. These properties permit enhanced interrogations of cell function, extended challenges to the regenerative capacities of discrete stem cell niches, and novel modeling of chronic degenerative diseases. Accordingly, we have created a series of bipartite transgenic reporter/effector resources to facilitate dissemination of NTR 2.0 to the research community.

Original languageEnglish (US)
Pages (from-to)205-215
Number of pages11
JournalNature Methods
Volume19
Issue number2
DOIs
StatePublished - Feb 2022

ASJC Scopus subject areas

  • Molecular Biology
  • Biochemistry
  • Biotechnology
  • Cell Biology

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