title: Kβ Valence to Core X-ray Emission Studies of Cu(I) Binding Proteins with Mixed Methionine - Histidine Coordination. Relevance to the Reactivity of the M- and H-sites of Peptidylglycine Monooxygenase.
abstract: Biological systems use copper as a redox center in many metalloproteins, where the role of the metal is to cycle between its +1 and +2 oxidation states. This chemistry requires the redox potential to be in a range that can stabilize both Cu(I) and Cu(II) states and often involves protein-derived ligand sets involving mixed histidine-methionine coordination that balance the preferences of both oxidation states. Transport proteins, on the other hand, utilize copper in the Cu(I) state and often contain sites comprised predominately of the cuprophilic residue methionine. The electronic factors that allow enzymes and transporters to balance their redox requirements are complex and are often elusive due to the dearth of spectroscopic probes of the Cu(I) state. Here we present the novel application of X-ray emission spectroscopy to copper proteins via a study of a series of mixed His-Met copper sites where the ligand set varies in a systematic way between the His3 and Met3 limits. The sites are derived from the wild-type peptidylglycine monooxygenase (PHM), two single-site variants which replicate each of its two copper sites (CuM-site and CuH-site), and the transporters CusF and CusB. Clear differences are observed in the Kβ2,5 region at the Met3 and His3 limits. CusB (Met3) has a distinct peak at 8978.4 eV with a broad shoulder at 8975.6 eV, whereas CuH (His3) has two well-resolved features: a more intense feature at 8974.8 eV and a second at 8977.2 eV. The mixed coordination sphere CusF (Met2His) and the PHM CuM variant (Met1His2) have very similar spectra consisting of two features at 8975.2 and 8977.8 eV. An analysis of DFT calculated spectra indicate that the intensity of the higher energy peak near 8978 eV is mediated by mixing of ligand-based orbitals into the Cu d(10) manifold, with S from Met providing more intensity by facilitating increased Cu p-d mixing. Furthermore, reaction of WT PHM with CO (an oxygen analogue) produced the M site CO complex, which showed a unique XES spectrum that could be computationally reproduced by including interactions between Cu(I) and the CO ligand. The study suggests that the valence-to-core (VtC) region can not only serve as a probe of ligand speciation but also offer insight into the coordination geometry, in a fashion similar to XAS pre-edges, and may be sufficiently sensitive to the coordination of exogenous ligands to be useful in the study of reaction mechanisms.
authors.affiliation: Max Planck Institute for Chemical Energy Conversion , Stiftstraße 34-36, D-45470 Mülheim an der Ruhr, Germany.
authors.forename: Vlad
authors.initials: V
authors.lastname: Martin-Diaconescu
authors.affiliation: Institute of Environmental Health, Oregon Health & Sciences University , 3181 SW Sam Jackson Park Road, Portland, Oregon 97239, United States.
authors.forename: Kelly N
authors.initials: KN
authors.lastname: Chacón
authors.affiliation: Max Planck Institute for Chemical Energy Conversion , Stiftstraße 34-36, D-45470 Mülheim an der Ruhr, Germany.
authors.forename: Mario Ulises
authors.initials: MU
authors.lastname: Delgado-Jaime
authors.affiliation: Stanford Synchrotron Radiation Lightsource , 2575 Sand Hill Road, Menlo Park, California 97025, United States.
authors.forename: Dimosthenis
authors.initials: D
authors.lastname: Sokaras
authors.affiliation: Stanford Synchrotron Radiation Lightsource , 2575 Sand Hill Road, Menlo Park, California 97025, United States.
authors.forename: Tsu-Chien
authors.initials: TC
authors.lastname: Weng
authors.affiliation: Max Planck Institute for Chemical Energy Conversion , Stiftstraße 34-36, D-45470 Mülheim an der Ruhr, Germany.|Department of Chemistry and Chemical Biology, Cornell University , Ithaca, New York 14853, United States.
authors.forename: Serena
authors.initials: S
authors.lastname: DeBeer
authors.affiliation: Institute of Environmental Health, Oregon Health & Sciences University , 3181 SW Sam Jackson Park Road, Portland, Oregon 97239, United States.
authors.forename: Ninian J
authors.initials: NJ
authors.lastname: Blackburn
mesh_terms: D000818:Animals; D001665:Binding Sites; D027682:Cation Transport Proteins / Q000737:chemistry* / Q000378:metabolism; D003300:Copper / Q000378:metabolism*; D000079922:Copper Transport Proteins; D004926:Escherichia coli / Q000737:chemistry* / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism; D006639:Histidine / Q000737:chemistry / Q000378:metabolism; D026901:Membrane Transport Proteins / Q000737:chemistry* / Q000378:metabolism; D008715:Methionine / Q000737:chemistry / Q000378:metabolism; D006899:Mixed Function Oxygenases / Q000737:chemistry* / Q000378:metabolism; D008958:Models, Molecular; D009097:Multienzyme Complexes / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D051381:Rats; D013052:Spectrometry, X-Ray Emission
chemical_list: D027682:Cation Transport Proteins; D000079922:Copper Transport Proteins; C556452:CusB protein, E coli; C506213:CusF protein, E coli; D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins; D009097:Multienzyme Complexes; D006639:Histidine; D003300:Copper; D008715:Methionine; D006899:Mixed Function Oxygenases; C035189:peptidylglycine monooxygenase