Mass spectrometric identification of intermediates in the O2-driven [4Fe-4S] to [2Fe-2S] cluster conversion in FNR

Jason C. Crack; Andrew J. Thomson; Nick E. Le Brun

doi:10.1073/pnas.1620987114

Mass spectrometric identification of intermediates in the O2-driven [4Fe-4S] to [2Fe-2S] cluster conversion in FNR

Jason C. Crack
, Andrew J. Thomson
, Nick E. Le Brun

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

52 Citations (Scopus)

36 Downloads (Pure)

Abstract

The iron-sulfur cluster containing protein FNR is the master regulator for the switch between anaerobic and aerobic respiration in Escherichia coli and many other bacteria. The [4Fe-4S] cluster functions as the sensory module, undergoing reaction with O2 that leads to conversion to a [2Fe-2S] form with loss of high affinity DNA-binding. Here we report studies of the FNR
cluster conversion reaction using time-resolved electrospray ionization mass spectrometry. The data provide new insight into the reaction, permitting the detection of cluster conversion intermediates and products, including a novel [3Fe-3S] cluster and persulfide coordinated [2Fe-2S] clusters ([2Fe-2S](S)n, where n = 1 or 2). Analysis of kinetic data revealed a branched mechanism in which cluster sulfide oxidation occurs in parallel with cluster conversion, and not as a subsequent, secondary reaction, to generate ([2Fe-2S](S)n species.
This methodology shows great potential for broad application to studies of protein cofactorsmall molecule interactions.

Original language	English
Pages (from-to)	E3215–E3223
Number of pages	9
Journal	Proceedings of the National Academy of Sciences of the United States of America (PNAS)
Volume	114
Issue number	6
Early online date	3 Apr 2017
DOIs	https://doi.org/10.1073/pnas.1620987114
Publication status	Published - 18 Apr 2017

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10.1073/pnas.1620987114

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Nick Le Brun
- N.Le-brunuea.acuk
- School of Chemistry, Pharmacy and Pharmacology - Professor of Biological Chemistry
- Centre for Molecular and Structural Biochemistry - Director
- Pharmacology and Biological Chemistry - Member
Person: Research Group Member, Academic, Teaching and Research

A new pathway for iron-sulphur cluster repair
Le Brun, N. (Principal Investigator), Crack, J. (Co-Investigator), Thomson, A. (Co-Investigator) & Cull, N. (Technician)
Biotechnology and Biological Sciences Research Council
30/05/14 → 29/05/17
Project: Research

Cite this

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title = "Mass spectrometric identification of intermediates in the O2-driven [4Fe-4S] to [2Fe-2S] cluster conversion in FNR",

abstract = "The iron-sulfur cluster containing protein FNR is the master regulator for the switch between anaerobic and aerobic respiration in Escherichia coli and many other bacteria. The [4Fe-4S] cluster functions as the sensory module, undergoing reaction with O2 that leads to conversion to a [2Fe-2S] form with loss of high affinity DNA-binding. Here we report studies of the FNR cluster conversion reaction using time-resolved electrospray ionization mass spectrometry. The data provide new insight into the reaction, permitting the detection of cluster conversion intermediates and products, including a novel [3Fe-3S] cluster and persulfide coordinated [2Fe-2S] clusters ([2Fe-2S](S)n, where n = 1 or 2). Analysis of kinetic data revealed a branched mechanism in which cluster sulfide oxidation occurs in parallel with cluster conversion, and not as a subsequent, secondary reaction, to generate ([2Fe-2S](S)n species. This methodology shows great potential for broad application to studies of protein cofactorsmall molecule interactions. ",

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doi = "10.1073/pnas.1620987114",

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Mass spectrometric identification of intermediates in the O2-driven [4Fe-4S] to [2Fe-2S] cluster conversion in FNR. / Crack, Jason C.; Thomson, Andrew J.; Le Brun, Nick E.
In: Proceedings of the National Academy of Sciences of the United States of America (PNAS), Vol. 114, No. 6, 18.04.2017, p. E3215–E3223.

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

TY - JOUR

T1 - Mass spectrometric identification of intermediates in the O2-driven [4Fe-4S] to [2Fe-2S] cluster conversion in FNR

AU - Crack, Jason C.

AU - Thomson, Andrew J.

AU - Le Brun, Nick E.

N1 - Early title: O2-sensing by [4Fe-4S] FNR: identification of intermediates during cluster conversion by mass spectrometry

PY - 2017/4/18

Y1 - 2017/4/18

N2 - The iron-sulfur cluster containing protein FNR is the master regulator for the switch between anaerobic and aerobic respiration in Escherichia coli and many other bacteria. The [4Fe-4S] cluster functions as the sensory module, undergoing reaction with O2 that leads to conversion to a [2Fe-2S] form with loss of high affinity DNA-binding. Here we report studies of the FNR cluster conversion reaction using time-resolved electrospray ionization mass spectrometry. The data provide new insight into the reaction, permitting the detection of cluster conversion intermediates and products, including a novel [3Fe-3S] cluster and persulfide coordinated [2Fe-2S] clusters ([2Fe-2S](S)n, where n = 1 or 2). Analysis of kinetic data revealed a branched mechanism in which cluster sulfide oxidation occurs in parallel with cluster conversion, and not as a subsequent, secondary reaction, to generate ([2Fe-2S](S)n species. This methodology shows great potential for broad application to studies of protein cofactorsmall molecule interactions.

AB - The iron-sulfur cluster containing protein FNR is the master regulator for the switch between anaerobic and aerobic respiration in Escherichia coli and many other bacteria. The [4Fe-4S] cluster functions as the sensory module, undergoing reaction with O2 that leads to conversion to a [2Fe-2S] form with loss of high affinity DNA-binding. Here we report studies of the FNR cluster conversion reaction using time-resolved electrospray ionization mass spectrometry. The data provide new insight into the reaction, permitting the detection of cluster conversion intermediates and products, including a novel [3Fe-3S] cluster and persulfide coordinated [2Fe-2S] clusters ([2Fe-2S](S)n, where n = 1 or 2). Analysis of kinetic data revealed a branched mechanism in which cluster sulfide oxidation occurs in parallel with cluster conversion, and not as a subsequent, secondary reaction, to generate ([2Fe-2S](S)n species. This methodology shows great potential for broad application to studies of protein cofactorsmall molecule interactions.

U2 - 10.1073/pnas.1620987114

DO - 10.1073/pnas.1620987114

M3 - Article

SN - 1091-6490

VL - 114

SP - E3215–E3223

JO - Proceedings of the National Academy of Sciences of the United States of America (PNAS)

JF - Proceedings of the National Academy of Sciences of the United States of America (PNAS)

IS - 6

ER -

Mass spectrometric identification of intermediates in the O2-driven [4Fe-4S] to [2Fe-2S] cluster conversion in FNR

Abstract

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A new pathway for iron-sulphur cluster repair

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