TitleRole of histidine-86 in the catalytic mechanism of ferredoxin:thioredoxin reductase
Publication TypeJournal Article
Year of Publication2009
AuthorsWalters, EM, Garcia-Serres, R, Naik, SG, Bourquin, F, Glauser, DA, Schurmann, P, Huynh, BH, Johnson, MK
Pagination1016 - 1024
Date Published2009///
ISBN Number0006-2960

Ferredoxin:thioredoxin reductase catalyzes the reduction of thioredoxins in plant chloroplasts using the [Fe2S2] ferredoxin as a one-electron donor and as such plays a central role in light regulation of oxygenic photosynthesis. The active-site comprises a [Fe4S4] cluster next to a redox-active disulfide that is cleaved in sequential one-electron steps and the combination of spectroscopic and crystallographic studies have revealed a catalytic mechanism involving novel site specific cluster chemistry in the oxidized, one-electron- and two-electron-reduced redox states. Histidine-86 has emerged as a potential proton donor/acceptor in the catalytic mechanism based on redox-related changes in the positioning of the imidazole ring during redox cycling and greatly decreased activity for the H86Y variant. Here we report on spectroscopic and redox characterization of the [Fe4S4] center in Synechocystis sp. PCC 6803 H86Y ferredoxin:thoredoxin reductase in the accessible redox states of both the as purified and N-ethylmaleimide-modified forms, using the combination of UV-visible absorption and variable-temperature magnetic circular dichroism, EPR, resonance Raman and Mossbauer spectroscopies. The results demonstrate that His86 is required for formation of the partially valence-localized [Fe4S4](2+) cluster that is the hallmark of two-electron-reduced intermediate. Taken together with the available structural data, the spectroscopic results indicate a functional role for His86 in protonation/deprotonation of the cluster-interacting thiol and anchoring the cluster interacting thiol in close proximity to the cluster in the two-electron-reduced intermediate

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