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Reconstitution of the [4Fe-4S] cluster in FNR and demonstration of the aerobic-anaerobic transcription switch in vitro. Biochem J 1996 Jun 15;316 ( Pt 3)(Pt 3):887-92

Date

06/15/1996

Pubmed ID

8670167

Pubmed Central ID

PMC1217433

DOI

10.1042/bj3160887

Scopus ID

2-s2.0-0029797082 (requires institutional sign-in at Scopus site)   162 Citations

Abstract

The FNR protein of Escherichia coli is a redox-responsive transcription regulator that activates and represses a family of genes required for anaerobic and aerobic metabolism. Reconstitution of wild-type FNR by anaerobic treatment with ferrous ions, cysteine and the NifS protein of Azotobacter vinelandii leads to the incorporation of two [4Fe-4S]2+ clusters per FNR dimer. The UV-visible spectrum of reconstituted FNR has a broad absorbance at 420 nm. The clusters are EPR silent under anaerobic conditions but are degraded to [3Fe-4S]+ by limited oxidation with air, and completely lost on prolonged air exposure. The association of FNR with the iron-sulphur clusters is confirmed by CD spectroscopy. Incorporation of the [4Fe-4S]2+ clusters increases site-specific DNA binding about 7-fold compared with apo-FNR. Anaerobic transcription activation and repression in vitro likewise depends on the presence of the iron-sulphur cluster, and its inactivation under aerobic conditions provides a demonstration in vitro of the FNR-mediated aerobic-anaerobic transcriptional switch.

Author List

Green J, Bennett B, Jordan P, Ralph ET, Thomson AJ, Guest JR

Author

Brian Bennett D.Phil. Professor and Chair in the Physics department at Marquette University




MESH terms used to index this publication - Major topics in bold

Aerobiosis
Anaerobiosis
Azotobacter vinelandii
Bacterial Proteins
Cysteine
DNA, Bacterial
Electron Spin Resonance Spectroscopy
Escherichia coli
Escherichia coli Proteins
Iron-Sulfur Proteins
Kinetics
Macromolecular Substances
Models, Structural
Promoter Regions, Genetic
Protein Conformation
Spectrophotometry
Transcription Factors
Transcription, Genetic