Characterization of Leader Peptide Binding during Catalysis by the Nisin Dehydratase NisB

Lindsay M. Repka; Kenton J. Hetrick; See Hyun Chee; Wilfred A. Van Der Donk

doi:10.1021/jacs.7b13506

Characterization of Leader Peptide Binding during Catalysis by the Nisin Dehydratase NisB

Lindsay M. Repka, Kenton J. Hetrick, See Hyun Chee, Wilfred A. Van Der Donk

Research output: Contribution to journal › Article › peer-review

Abstract

The dehydratase NisB performs stepwise tRNA^Glu-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 tRNA^Glu binding to NisB that enables dehydration.

Original language	English (US)
Pages (from-to)	4200-4203
Number of pages	4
Journal	Journal of the American Chemical Society
Volume	140
Issue number	12
DOIs	https://doi.org/10.1021/jacs.7b13506
State	Published - Mar 28 2018

ASJC Scopus subject areas

Catalysis
General Chemistry
Biochemistry
Colloid and Surface Chemistry

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10.1021/jacs.7b13506

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title = "Characterization of Leader Peptide Binding during Catalysis by the Nisin Dehydratase NisB",

abstract = "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.",

author = "Repka, {Lindsay M.} and Hetrick, {Kenton J.} and Chee, {See Hyun} and {Van Der Donk}, {Wilfred A.}",

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T1 - Characterization of Leader Peptide Binding during Catalysis by the Nisin Dehydratase NisB

AU - Repka, Lindsay M.

AU - Hetrick, Kenton J.

AU - Chee, See Hyun

AU - Van Der Donk, Wilfred A.

PY - 2018/3/28

Y1 - 2018/3/28

N2 - 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.

AB - 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.

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Characterization of Leader Peptide Binding during Catalysis by the Nisin Dehydratase NisB

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