title: Role of histidine-86 in the catalytic mechanism of ferredoxin:thioredoxin reductase.
abstract: 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 Mössbauer 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.
authors.affiliation: Department of Chemistry and Center for Metalloenzyme Studies, University of Georgia, Athens, Georgia 30602, USA.
authors.forename: Elizabeth M
authors.initials: EM
authors.lastname: Walters
authors.forename: Ricardo
authors.initials: R
authors.lastname: Garcia-Serres
authors.forename: Sunil G
authors.initials: SG
authors.lastname: Naik
authors.forename: Florence
authors.initials: F
authors.lastname: Bourquin
authors.forename: Dominique A
authors.initials: DA
authors.lastname: Glauser
authors.forename: Peter
authors.initials: P
authors.lastname: Schürmann
authors.forename: Boi Hanh
authors.initials: BH
authors.lastname: Huynh
authors.forename: Michael K
authors.initials: MK
authors.lastname: Johnson
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism / Q000502:physiology; D002384:Catalysis; D004583:Electrons; D005288:Ferredoxins / Q000737:chemistry* / Q000502:physiology; D006639:Histidine / Q000737:chemistry* / Q000235:genetics; D007506:Iron-Sulfur Proteins / Q000737:chemistry; D010084:Oxidation-Reduction; D011522:Protons; D013438:Sulfhydryl Compounds / Q000737:chemistry; D046939:Synechocystis / Q000201:enzymology / Q000378:metabolism; D013880:Thioredoxin-Disulfide Reductase / Q000737:chemistry* / Q000502:physiology
chemical_list: D001426:Bacterial Proteins; D005288:Ferredoxins; D007506:Iron-Sulfur Proteins; D011522:Protons; D013438:Sulfhydryl Compounds; D006639:Histidine; D013880:Thioredoxin-Disulfide Reductase