| Autor: |
Bowman HE; Department of Chemistry, University of Wisconsin-Madison, 1101 University Ave., Madison, WI, 53706, USA., Dent MR; Department of Chemistry, University of Wisconsin-Madison, 1101 University Ave., Madison, WI, 53706, USA., Burstyn JN; Department of Chemistry, University of Wisconsin-Madison, 1101 University Ave., Madison, WI, 53706, USA. burstyn@chem.wisc.edu. |
| Jazyk: |
angličtina |
| Zdroj: |
Journal of biological inorganic chemistry : JBIC : a publication of the Society of Biological Inorganic Chemistry [J Biol Inorg Chem] 2016 Jul; Vol. 21 (4), pp. 559-69. Date of Electronic Publication: 2016 Jun 09. |
| DOI: |
10.1007/s00775-016-1368-5 |
| Abstrakt: |
Both Met(104) and Met(105) are involved, either directly or indirectly, in the redox mediated ligand switch of the heme-dependent transcription factor, RcoM-1. Recent studies of Burkholderia xenovorans RcoM identified Cys(94) as the thiolate ligand in the Fe(III) state of the heme cofactor. Upon reduction, a neutral donor replaces Cys(94) trans to His(74). Homology modelling implicated either Met(104) or Met(105) as the possible ligand in the Fe(II) state. We spectroscopically compared wild type (WT) RcoM-1 to three Met-to-Leu variants (M104L, M105L, and M104L/M105L) to identify which Met residue acts as the ligand. All proteins were isolated as admixtures of Fe(III) and Fe(II)-CO heme; oxidation by ferricyanide enables study of homogeneous oxidation and coordination states. Met(104) is the CO-replaceable Fe(II) heme ligand. The magnetic circular dichroism (MCD) spectrum of Fe(II) M105L resembled WT. M104L and M104L/M105L, however, showed spectra arising from the formation of a high-spin, five-coordinate species indicating the loss of the ligand. The electron paramagnetic resonance (EPR) spectra of WT Fe(III) RcoM-1, oxidized Fe(III) M104L, and as-isolated M105L exhibited narrow, rhombic low-spin signals typical of thiolate-bound hemes. In contrast, oxidized Fe(III) M105L and oxidized Fe(III) M104L/M105L revealed a broad, rhombic low-spin, six-coordinate signal indicative of replacement of the thiolate by a neutral ligand. Thus, we conclude that Met(105) is important to the stability of the Fe(III) heme pocket during oxidation. |
| Databáze: |
MEDLINE |
| Externí odkaz: |
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