Heme binding to the second, lower-affinity site of the global iron regulator Irr from Rhizobium leguminosarum promotes oligomerization

White, Gaye F., Singleton, Chloe, Todd, Jonathan D., Cheesman, Myles R., Johnston, Andrew W. B. and Le Brun, Nick E. ORCID: https://orcid.org/0000-0001-9780-4061 (2011) Heme binding to the second, lower-affinity site of the global iron regulator Irr from Rhizobium leguminosarum promotes oligomerization. FEBS Journal, 278 (12). pp. 2011-2021. ISSN 1742-4658

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Abstract

The iron responsive regulator Irr is found in a wide range of a-proteobacteria, where it regulates many genes in response to the essential but toxic metal iron. Unlike Fur, the transcriptional regulator that is used for iron homeostasis by almost all other bacterial lineages, Irr does not sense Fe2+ directly, but, rather, interacts with a physiologically important form of iron, namely heme. Recent studies of Irr from the N2-fixing symbiont Rhizobium leguminosarum (IrrRl) showed that it binds heme with submicromolar affinity at a His-Xxx-His (HxH) motif. This caused the protein to dissociate from its cognate DNA regulatory iron control element box sequences, thus allowing expression of its target genes under iron-replete conditions. In the present study, we report new insights into the mechanisms and consequences of heme binding to Irr. In addition to the HxH motif, Irr binds heme at a second, lower-affinity site. Spectroscopic studies of wild-type Irr and His variants show that His46 and probably His66 are involved in coordinating heme in a low-spin state at this second site. By contrast to the well-studied Irr from Bradyrhizobium japonicum, neither heme site of IrrRl stabilizes ferrous heme. Furthermore, we show that heme-free IrrRl exists as a mixture of dimeric and larger, likely hexameric, forms and that heme binding promotes IrrRl oligomerization. Bioanalytical studies of IrrRl variants showed that this property is not dependent on the HxH motif but is associated with heme binding at the second site.

Item Type:	Article
Faculty \ School:	Faculty of Science > School of Biological Sciences Faculty of Science > School of Chemistry
UEA Research Groups:	Faculty of Science > Research Groups > Molecular Microbiology Faculty of Science > Research Groups > Chemistry of Light and Energy Faculty of Science > Research Groups > Chemistry of Life Processes Faculty of Science > Research Groups > Biophysical Chemistry (former - to 2017) Faculty of Science > Research Centres > Centre for Molecular and Structural Biochemistry Faculty of Science > Research Groups > Centre for Ocean and Atmospheric Sciences
Depositing User:	Users 2731 not found.
Date Deposited:	09 Jan 2012 13:15
Last Modified:	09 Mar 2024 20:30
URI:	https://ueaeprints.uea.ac.uk/id/eprint/35962
DOI:	10.1111/j.1742-4658.2011.08117.x

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