Cell Chemical Biology
Volume 26, Issue 6, 20 June 2019, Pages 878-884.e8
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Brief Communication
Late-Stage Conversion of Diphenylphosphonate to Fluorophosphonate Probes for the Investigation of Serine Hydrolases

https://doi.org/10.1016/j.chembiol.2019.02.020Get rights and content
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Highlights

  • Facile synthesis of structurally complex, reactive peptide methylfluorophosphonates

  • Diverse functional group tolerance without protecting groups

  • Mechanistic studies support the intermediacy of a difluorophosphonate

  • Specific inactivation of the nocardicin biosynthetic thioesterase exemplified

Summary

Diphenylphosphonates (DPPs) have been used for 50 years to inactivate serine hydrolases, creating adducts representative of tetrahedral intermediates of this important class of enzymes. Failure to react at active site serine residues, however, can thwart their usefulness. Here, we describe a facile route and allied mechanistic studies to highly reactive, structurally complex organofluorophosphonates (FPs) by direct fluorinative hydrolysis of DPPs. Advantages over current preparations of FPs are exemplified by the synthesis of a β-lactam-containing peptide substrate analog capable of modifying the C-terminal, dual-function thioesterase involved in nocardicin A biosynthesis. Although this serine hydrolase was found to resist modification by classic DPP inhibitors, active site selective phosphonylation by the corresponding FP occurs rapidly to form a stable adduct. This simple, late-stage method enables the ready preparation of FP probes that retain important structural motifs of native substrates, thus promoting efforts in mechanistic enzymology by accessing biologically relevant enzyme-inhibitor co-structures.

Keywords

alkylfluorophosphonate synthesis
diphenylphosphonate synthesis
serine hydrolase
mechanism-based inhibitor
enzyme inactivation

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2

Present address: Tri-Institutional MD-PhD Program, Weill Cornell Medical College, 1300 York Avenue, New York, NY 10065, USA

3

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