{"id":635,"date":"2026-04-15T13:27:02","date_gmt":"2026-04-15T13:27:02","guid":{"rendered":"https:\/\/citations.tools.bio-logic.fr\/?p=635"},"modified":"2026-04-15T13:27:02","modified_gmt":"2026-04-15T13:27:02","slug":"evidence-for-a-long-lived-cu-coupled-and-oxygen-inert-disulfide-radical-anion-in-the-assembly-of-metallothionein-3-cui4-thiolate-cluster","status":"publish","type":"post","link":"https:\/\/citations.tools.bio-logic.fr\/?p=635","title":{"rendered":"Evidence for a Long-Lived, Cu-Coupled and Oxygen-Inert Disulfide Radical Anion in the Assembly of Metallothionein-3 Cu(I)<sub>4<\/sub>-Thiolate Cluster"},"content":{"rendered":"<h4>DOI:<\/h4>\n<p><a href=\"https:\/\/doi.org\/10.1021\/jacs.1c03984\" target=\"_blank\" rel=\"noopener\">10.1021\/jacs.1c03984<\/a><\/p>\n<h4>Authors:<\/h4>\n<p>Jenifer S. Calvo, Rhiza Lyne E. Villones, Nicholas J. York, Ewelina Stefaniak, Grace E. Hamilton, Allison L. Stelling, Wojciech Bal, Brad S. Pierce, Gabriele Meloni<\/p>\n<h4>Abstract:<\/h4>\n<p>The human copper-binding protein metallothionein-3 (MT-3) can reduce Cu(II) to Cu(I) and form a polynuclear Cu(I)<sub>4<\/sub>-Cys<sub>5-6<\/sub> cluster concomitant with intramolecular disulfide bonds formation, but the cluster is unusually inert toward O<sub>2<\/sub> and redox-cycling. We utilized a combined array of rapid-mixing spectroscopic techniques to identify and characterize the transient radical intermediates formed in the reaction between Zn<sub>7<\/sub>MT-3 and Cu(II) to form Cu(I)<sub>4<\/sub>Zn(II)<sub>4<\/sub>MT-3. Stopped-flow electronic absorption spectroscopy reveals the rapid formation of transient species with absorption centered at 430-450 nm and consistent with the generation of disulfide radical anions (DRAs) upon reduction of Cu(II) by MT-3 cysteine thiolates. These DRAs are oxygen-stable and unusually long-lived, with lifetimes in the seconds regime. Subsequent DRAs reduction by Cu(II) leads to the formation of a redox-inert Cu(I)<sub>4<\/sub>-Cys<sub>5<\/sub> cluster with short Cu-Cu distances (&lt;2.8 \u00c5), as revealed by low-temperature (77 K) luminescence spectroscopy. Rapid freeze-quench Raman and electron paramagnetic resonance (EPR) spectroscopy characterization of the intermediates confirmed the DRA nature of the sulfur-centered radicals and their subsequent oxidation to disulfide bonds upon Cu(II) reduction, generating the final Cu(I)<sub>4<\/sub>-thiolate cluster. EPR simulation analysis of the radical <i>g<\/i>&#8211; and <i>A<\/i>-values indicate that the DRAs are directly coupled to Cu(I), potentially explaining the observed DRA stability in the presence of O<sub>2<\/sub>. We thus provide evidence that the MT-3 Cu(I)<sub>4<\/sub>-Cys<sub>5<\/sub> cluster assembly process involves the controlled formation of novel long-lived, copper-coupled, and oxygen-stable disulfide radical anion transient intermediates.<\/p>\n","protected":false},"excerpt":{"rendered":"<p>DOI: 10.1021\/jacs.1c03984 Authors: Jenifer S. Calvo, Rhiza Lyne E. Villones, Nicholas J. York, Ewelina Stefaniak, Grace E. Hamilton, Allison L. Stelling, Wojciech Bal, Brad S. Pierce, Gabriele Meloni Abstract: The human copper-binding protein metallothionein-3 (MT-3) can reduce Cu(II) to Cu(I) and form a polynuclear Cu(I)4-Cys5-6 cluster concomitant with intramolecular disulfide bonds formation, but the cluster [&hellip;]<\/p>\n","protected":false},"author":1,"featured_media":0,"comment_status":"open","ping_status":"open","sticky":false,"template":"","format":"standard","meta":{"footnotes":""},"categories":[5],"tags":[],"class_list":["post-635","post","type-post","status-publish","format-standard","hentry","category-freeze-quench"],"_links":{"self":[{"href":"https:\/\/citations.tools.bio-logic.fr\/index.php?rest_route=\/wp\/v2\/posts\/635","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/citations.tools.bio-logic.fr\/index.php?rest_route=\/wp\/v2\/posts"}],"about":[{"href":"https:\/\/citations.tools.bio-logic.fr\/index.php?rest_route=\/wp\/v2\/types\/post"}],"author":[{"embeddable":true,"href":"https:\/\/citations.tools.bio-logic.fr\/index.php?rest_route=\/wp\/v2\/users\/1"}],"replies":[{"embeddable":true,"href":"https:\/\/citations.tools.bio-logic.fr\/index.php?rest_route=%2Fwp%2Fv2%2Fcomments&post=635"}],"version-history":[{"count":0,"href":"https:\/\/citations.tools.bio-logic.fr\/index.php?rest_route=\/wp\/v2\/posts\/635\/revisions"}],"wp:attachment":[{"href":"https:\/\/citations.tools.bio-logic.fr\/index.php?rest_route=%2Fwp%2Fv2%2Fmedia&parent=635"}],"wp:term":[{"taxonomy":"category","embeddable":true,"href":"https:\/\/citations.tools.bio-logic.fr\/index.php?rest_route=%2Fwp%2Fv2%2Fcategories&post=635"},{"taxonomy":"post_tag","embeddable":true,"href":"https:\/\/citations.tools.bio-logic.fr\/index.php?rest_route=%2Fwp%2Fv2%2Ftags&post=635"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}