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Characterization of Leader Peptide Binding During Catalysis by the Nisin Dehydratase NisB
- Repka, Lindsay M., Hetrick, Kenton J., Chee, See Hyun, van der Donk, Wilfred A.
- Journal of the American Chemical Society 2018 v.140 no.12 pp. 4200-4203
- antimicrobial peptides, binding sites, biosynthesis, catalytic activity, chemical bonding, nisin, signal peptide
- The dehydratase NisB performs stepwise tRNAᴳˡᵘ-dependent glutamylation of Ser/Thr residues and subsequent glutamate elimination to effect eight dehydrations in the biosynthesis of the antibacterial peptide nisin. Its substrate, NisA, bears a C-terminal core peptide that is modified and an N-terminal leader peptide (LP) that is not modified but that is required for efficient dehydration. To elucidate the mechanism of LP-NisB interactions during dehydration, we engineered a disulfide that covalently links the NisA LP to NisB. The enzyme fully dehydrated tethered NisA, confirming the functional LP binding site and supporting a mechanism where NisB uses a single LP binding site for glutamylation and elimination. We also show an order of NisA and tRNAᴳˡᵘ binding to NisB that enables dehydration.