title: Tyrosine-Coordinated P-Cluster in G. diazotrophicus Nitrogenase: Evidence for the Importance of O-Based Ligands in Conformationally Gated Electron Transfer.
abstract: The P-cluster is a unique iron-sulfur center that likely functions as a dynamic electron (e(-)) relay site between the Fe-protein and the catalytic FeMo-cofactor in nitrogenase. The P-cluster has been shown to undergo large conformational changes upon 2-e(-) oxidation which entail the coordination of two of the Fe centers to a Ser side chain and a backbone amide N, respectively. Yet, how and if this 2-e(-) oxidized state (P(OX)) is involved in catalysis by nitrogenase is not well established. Here, we present the crystal structures of reduced and oxidized MoFe-protein (MoFeP) from Gluconacetobacter diazotrophicus (Gd), which natively possesses an Ala residue in the position of the Ser ligand to the P-cluster. While reduced Gd-MoFeP is structurally identical to previously characterized counterparts around the FeMo-cofactor, oxidized Gd-MoFeP features an unusual Tyr coordination to its P-cluster along with ligation by a backbone amide nitrogen. EPR analysis of the oxidized Gd-MoFeP P-cluster confirmed that it is a 2-e(-) oxidized, integer-spin species. Importantly, we have found that the sequence positions corresponding to the Ser and Tyr ligands are almost completely covariant among Group I nitrogenases. These findings strongly support the possibility that the P(OX) state is functionally relevant in nitrogenase catalysis and that a hard, O-based anionic ligand serves to stabilize this state in a switchable fashion.
authors.affiliation: Department of Chemistry and Biochemistry, University of California, San Diego , 9500 Gilman Drive, La Jolla, California 92093-0356, United States.
authors.forename: Cedric P
authors.initials: CP
authors.lastname: Owens
authors.affiliation: Department of Chemistry and Biochemistry, University of California, San Diego , 9500 Gilman Drive, La Jolla, California 92093-0356, United States.
authors.forename: Faith E H
authors.initials: FE
authors.lastname: Katz
authors.affiliation: Department of Chemistry and Biochemistry, University of California, San Diego , 9500 Gilman Drive, La Jolla, California 92093-0356, United States.
authors.forename: Cole H
authors.initials: CH
authors.lastname: Carter
authors.affiliation: Department of Chemistry, University of California , Irvine, 1102 Natural Science II, Irvine, California 92697, United States.
authors.forename: Victoria F
authors.initials: VF
authors.lastname: Oswald
authors.affiliation: Department of Chemistry and Biochemistry, University of California, San Diego , 9500 Gilman Drive, La Jolla, California 92093-0356, United States.
authors.forename: F Akif
authors.initials: FA
authors.lastname: Tezcan
mesh_terms: D000409:Alanine / Q000737:chemistry; D001426:Bacterial Proteins / Q000737:chemistry; D001665:Binding Sites; D002384:Catalysis; D004578:Electron Spin Resonance Spectroscopy; D004579:Electron Transport; D004583:Electrons; D041821:Gluconacetobacter / Q000201:enzymology*; D007506:Iron-Sulfur Proteins / Q000737:chemistry; D008024:Ligands; D008983:Molybdoferredoxin / Q000378:metabolism; D009591:Nitrogenase / Q000737:chemistry*; D010084:Oxidation-Reduction; D010100:Oxygen / Q000737:chemistry; D011487:Protein Conformation; D014443:Tyrosine / Q000737:chemistry*
chemical_list: D001426:Bacterial Proteins; D007506:Iron-Sulfur Proteins; D008024:Ligands; D008983:Molybdoferredoxin; D014443:Tyrosine; D009591:Nitrogenase; D000409:Alanine; D010100:Oxygen