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. 2017 Jul 21;7:6179. doi: 10.1038/s41598-017-05268-2

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© The Author(s) 2017

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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EPR analysis of Lcp_K30. Continuous-wave EPR spectra were recorded for Lcp_K30 as isolated (blue), Lcp_K30 with imidazole (1 mM, red) and the Lcp_K30 Lys167Ala variant (grey). Three discernible signals are a broad near-axial signal originating from a six-coordinate low-spin haem, most likely with Lys167 coordinating the haem iron. A signal at g~6 represents a second population of five-coordinate high-spin haem. Added imidazole binds to the haem group, replacing Lys167 and shifting the protein to a low-spin state. The high-spin signal disappears, and the g-values of the low-spin signal shift to 1.52, 2.27 and 2.97, representing the binding of imidazole, with the total population of paramagnetic Fe³⁺ being reduced.