Details

Autor(en) / Beteiligte

• Bates, Donna M.
• Popescu, Codrina V.
• Khoroshilova, Natalia
• Vogt, Kevin
• Beinert, Helmut
• Münck, Eckard
• Kiley, Patricia J.

Titel

Substitution of Leucine 28 with Histidine in theEscherichia coli Transcription Factor FNR Results in Increased Stability of the [4Fe-4S]2+ Cluster to Oxygen

Ist Teil von

• The Journal of biological chemistry, 2000-03, Vol.275 (9), p.6234-6240

Ort / Verlag

Elsevier Inc

Erscheinungsjahr

2000

Quelle

Alma/SFX Local Collection

Beschreibungen/Notizen

• To understand the role of the [4Fe-4S]2+ cluster in controlling the activity of the Escherichia coli transcription factor FNR (fumarate nitrate reduction) during changes in O2 availability, we have characterized a mutant FNR protein containing a substitution of Leu-28 with His (FNR-L28H) which, unlike its wild type (WT) counterpart, is functional under aerobic growth conditions. The His-28 substitution appears to stabilize the [4Fe-4S]2+ cluster of FNR-L28H in the presence of O2 because air-exposed FNR-L28H did not undergo the rapid [4Fe-4S]2+ to [2Fe-2S]2+ cluster conversion or concomitant loss in site-specific DNA binding and dimerization, which are characteristic of WT-FNR under these conditions. This increased cluster stability was not a result of His-28 replacing the WT-FNR cluster ligands because substitution of any of these four Cys residues (cysteine 20, 23, 29, or 122) with Ser resulted in [4Fe-4S]2+ cluster-deficient preparations of FNR-L28H. The Mössbauer spectra of FNR-L28H indicated that the coordination environment of the [4Fe-4S]2+ cluster did not differ from that of WT-FNR. Whole cell Mössbauer spectroscopy showed that aerobically grown cells overexpressing FNR-L28H had levels of the FNR species containing the [4Fe-4S]2+ cluster similar to those of cells grown under anaerobic conditions. Thus, the increase in cluster stability is sufficient to allow accumulation of the [4Fe-4S]2+cluster form of FNR-L28H under aerobic conditions and provides a reasonable explanation for why this mutant protein is functional under aerobic growth conditions. From these results, we present a model to explain how WT-FNR is normally inactivated under aerobic growth conditions.