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. 2006 Feb 8;128(5):1420-1.
doi: 10.1021/ja057203d.

The dehydratase activity of lacticin 481 synthetase is highly processive

Leah M Miller  1 , Champak Chatterjee, Wilfred A van der Donk, Neil L Kelleher
Affiliations

The dehydratase activity of lacticin 481 synthetase is highly processive

Leah M Miller et al. J Am Chem Soc. .

Abstract

Lacticin 481 synthetase (LctM) is a bifunctional enzyme that undertakes dehydration and cyclization in the structural region of the pre-lacticin peptide (LctA) to introduce three thioether rings and one dehydrobutyrine residue. The order and timing of these events has been investigated employing high-resolution ESI-FTMS-based tandem MS/MS techniques and chemical derivatization. LctM demonstrates highly processive behavior as seen by MS analysis of the reaction course of dehydration. Furthermore, cyclization is not tightly coupled to dehydration and follows at a later stage of the enzymatic reaction.

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Figures

Figure 1
Figure 1
A) LanM catalyzed formation of (Me)Lan. B) Time dependent ESI/FT mass spectra of the assay of His6-LctA with LctM. * = oxidized products M+(O).
Figure 2
Figure 2
Assay product of LctM with His6-LctA at 1:1 enzyme to substrate ratio. The 3 s time-point exhibits the presence of all possible dehydrated forms of LctA (7+ charge state).
Figure 3
Figure 3
ESI/FT MS/MS interrogation of thioether formation in His6-LctA during LctM catalysis. A) Expected cyanylation products. B) Cyanylation of the assay products after 40 min. Colored asterisks depict oxidation products, M+(O), of the identically colored parent ions. C) MS/MS analysis of the LctA-3 H2O species from Figure 1B (40 min). D) Fragment ion map of the LctA-3 H2O species.
Scheme 1
Scheme 1
See this image and copyright information in PMC

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