Huguenin-Dezot, N. et al. Nature 565, 112–117 (2019).
Many enzymatic reactions, including those of serine hydrolases and cysteine proteases, proceed via the formation of an unstable acyl-enzyme intermediate. Because these intermediates exist in fleeting states, they are hard to study structurally, which prevents mechanistic analysis. Huguenin-Dezot et al. describe a trapping strategy for stabilizing such intermediate enzyme states. By using genetic code expansion methods, they incorporate the unnatural amino acid DAP (2,3-diaminopropionic acid) into an enzyme of interest, in place of the catalytic serine or cysteine residue. DAP acts as a nucleophile, forming an amide bond with the enzyme substrate, and effectively trapping the acyl-enzyme intermediate. The trapped intermediate states are more amenable to structure determination, such as by X-ray crystallography. The authors applied this approach to study the mechanism by which valinomycin synthetase oligomerizes and cyclizes its tetradepsipetidyl substrate to produce the natural product valinomycin.
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Doerr, A. A trap for intermediate enzyme complexes. Nat Methods 16, 217 (2019). https://doi.org/10.1038/s41592-019-0339-5
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DOI: https://doi.org/10.1038/s41592-019-0339-5