{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,4,20]],"date-time":"2026-04-20T19:36:32Z","timestamp":1776713792349,"version":"3.51.2"},"reference-count":77,"publisher":"MDPI AG","issue":"2","license":[{"start":{"date-parts":[[2024,1,20]],"date-time":"2024-01-20T00:00:00Z","timestamp":1705708800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100000780","name":"European Union\u2019s Horizon 2020 research and innovation programme","doi-asserted-by":"publisher","award":["813209"],"award-info":[{"award-number":["813209"]}],"id":[{"id":"10.13039\/501100000780","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100000780","name":"European Union\u2019s Horizon 2020 research and innovation programme","doi-asserted-by":"publisher","award":["PID2021-127287NB-I00"],"award-info":[{"award-number":["PID2021-127287NB-I00"]}],"id":[{"id":"10.13039\/501100000780","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100004837","name":"Spanish Ministry of Science and Innovation","doi-asserted-by":"publisher","award":["813209"],"award-info":[{"award-number":["813209"]}],"id":[{"id":"10.13039\/501100004837","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100004837","name":"Spanish Ministry of Science and Innovation","doi-asserted-by":"publisher","award":["PID2021-127287NB-I00"],"award-info":[{"award-number":["PID2021-127287NB-I00"]}],"id":[{"id":"10.13039\/501100004837","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Molecules"],"abstract":"<jats:p>The cytochrome P450 family consists of ubiquitous monooxygenases with the potential to perform a wide variety of catalytic applications. Among the members of this family, CYP116B5hd shows a very prominent resistance to peracid damage, a property that makes it a promising tool for fine chemical synthesis using the peroxide shunt. In this meticulous study, we use hyperfine spectroscopy with a multifrequency approach (X- and Q-band) to characterize in detail the electronic structure of the heme iron of CYP116B5hd in the resting state, which provides structural details about its active site. The hyperfine dipole\u2013dipole interaction between the electron and proton nuclear spins allows for the locating of two different protons from the coordinated water and a beta proton from the cysteine axial ligand of heme iron with respect to the magnetic axes centered on the iron. Additionally, since new anti-cancer therapies target the inhibition of P450s, here we use the CYP116B5hd system\u2014imidazole as a model for studying cytochrome P450 inhibition by an azo compound. The effects of the inhibition of protein by imidazole in the active-site geometry and electron spin distribution are presented. The binding of imidazole to CYP116B5hd results in an imidazole\u2013nitrogen axial coordination and a low-spin heme FeIII. HYSCORE experiments were used to detect the hyperfine interactions. The combined interpretation of the gyromagnetic tensor and the hyperfine and quadrupole tensors of magnetic nuclei coupled to the iron electron spin allowed us to obtain a precise picture of the active-site geometry, including the orientation of the semi-occupied orbitals and magnetic axes, which coincide with the porphyrin N-Fe-N axes. The electronic structure of the iron does not seem to be affected by imidazole binding. Two different possible coordination geometries of the axial imidazole were observed. The angles between gx (coinciding with one of the N-Fe-N axes) and the projection of the imidazole plane on the heme were determined to be \u221260\u00b0 and \u221225\u00b0 for each of the two possibilities via measurement of the hyperfine structure of the axially coordinated 14N.<\/jats:p>","DOI":"10.3390\/molecules29020518","type":"journal-article","created":{"date-parts":[[2024,1,22]],"date-time":"2024-01-22T11:36:41Z","timestamp":1705923401000},"page":"518","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":4,"title":["Heme Spin Distribution in the Substrate-Free and Inhibited Novel CYP116B5hd: A Multifrequency Hyperfine Sublevel Correlation (HYSCORE) Study"],"prefix":"10.3390","volume":"29","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-5431-2371","authenticated-orcid":false,"given":"Antonino","family":"Famulari","sequence":"first","affiliation":[{"name":"Departamento de F\u00edsica de la Materia Condensada, Universidad de Zaragoza, C\/Pedro Cerbuna 12, 50009 Zaragoza, Spain"},{"name":"Department of Chemistry, University of Turin, Via Giuria 9, 10125 Torino, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4945-8904","authenticated-orcid":false,"given":"Danilo","family":"Correddu","sequence":"additional","affiliation":[{"name":"Department of Life Sciences and Systems Biology, University of Turin, Via Accademia Albertina 13, 10123 Torino, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4169-2635","authenticated-orcid":false,"given":"Giovanna","family":"Di Nardo","sequence":"additional","affiliation":[{"name":"Department of Life Sciences and Systems Biology, University of Turin, Via Accademia Albertina 13, 10123 Torino, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6559-276X","authenticated-orcid":false,"given":"Gianfranco","family":"Gilardi","sequence":"additional","affiliation":[{"name":"Department of Life Sciences and Systems Biology, University of Turin, Via Accademia Albertina 13, 10123 Torino, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7874-2846","authenticated-orcid":false,"given":"George","family":"Mitrikas","sequence":"additional","affiliation":[{"name":"Institute of Nanoscience and Nanotechnology, NCSR Demokritos, 15341 Athens, Greece"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Mario","family":"Chiesa","sequence":"additional","affiliation":[{"name":"Department of Chemistry, University of Turin, Via Giuria 9, 10125 Torino, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1827-1250","authenticated-orcid":false,"given":"In\u00e9s","family":"Garc\u00eda-Rubio","sequence":"additional","affiliation":[{"name":"Departamento de F\u00edsica de la Materia Condensada, Universidad de Zaragoza, C\/Pedro Cerbuna 12, 50009 Zaragoza, Spain"},{"name":"Instituto de Nanociencia y Materiales de Arag\u00f3n (INMA), CSIC-Universidad de Zaragoza, 50009 Zaragoza, Spain"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2024,1,20]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","unstructured":"Ortiz de Montellano, P.R. 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