C.M.Czekster
et al.
(2017).
Characterization of a dual function macrocyclase enables design and use of efficient macrocyclization substrates.
Nat Commun,
8,
1045.
PubMed id: 29051530
Peptide macrocycles are promising therapeutic molecules because they are
protease resistant, structurally rigid, membrane permeable, and capable of
modulating protein-protein interactions. Here, we report the characterization of
the dual function macrocyclase-peptidase enzyme involved in the biosynthesis of
the highly toxic amanitin toxin family of macrocycles. The enzyme first removes
10 residues from the N-terminus of a 35-residue substrate. Conformational
trapping of the 25 amino-acid peptide forces the enzyme to release this
intermediate rather than proceed to macrocyclization. The enzyme rebinds the 25
amino-acid peptide in a different conformation and catalyzes macrocyclization of
the N-terminal eight residues. Structures of the enzyme bound to both substrates
and biophysical analysis characterize the different binding modes rationalizing
the mechanism. Using these insights simpler substrates with only five C-terminal
residues were designed, allowing the enzyme to be more effectively exploited in
biotechnology.
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