title: Chemically distinct Ni sites in the A-cluster in subunit beta of the acetyl-CoA decarbonylase/synthase complex from Methanosarcina thermophila: Ni L-edge absorption and X-ray magnetic circular dichroism analyses.
abstract: The 5-subunit-containing acetyl-CoA decarbonylase/synthase (ACDS) complex plays an important role in methanogenic Archaea that convert acetate to methane, by catalyzing the central reaction of acetate C-C bond cleavage in which acetyl-CoA serves as the acetyl donor substrate reacting at the ACDS beta subunit active site. The properties of Ni in the active site A-cluster in the ACDS beta subunit from Methanosarcina thermophila were investigated. A recombinant, C-terminally truncated form of the beta subunit was employed, which mimics the native subunit previously isolated from the ACDS complex, and contains an A-cluster composed of an [Fe(4)S(4)] center bridged to a binuclear Ni-Ni site. The electronic structures of these two Ni were studied using L-edge absorption and X-ray magnetic circular dichroism (XMCD) spectroscopy. The L-edge absorption data provided evidence for two distinct Ni species in the as-isolated enzyme, one with low-spin Ni(II) and the other with high-spin Ni(II). XMCD spectroscopy confirmed that the species producing the high-spin signal was paramagnetic. Upon treatment with Ti(3+) citrate, an additional Ni species emerged, which was assigned to Ni(I). By contrast, CO treatment of the reduced enzyme converted nearly all of the Ni in the sample to low-spin Ni(II). The results implicate reaction of a high-spin tetrahedral Ni site with CO to form an enzyme-CO adduct transformed to a low-spin Ni(II) state. These findings are discussed in relation to the mechanism of C-C bond activation, in connection with the model of the beta subunit A-cluster developed from companion Ni and Fe K edge, XANES, and EXAFS studies.
authors.affiliation: Physical Biosciences Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720, USA.
authors.forename: Tobias
authors.initials: T
authors.lastname: Funk
authors.forename: Weiwei
authors.initials: W
authors.lastname: Gu
authors.forename: Stephan
authors.initials: S
authors.lastname: Friedrich
authors.forename: Hongxin
authors.initials: H
authors.lastname: Wang
authors.forename: Simonida
authors.initials: S
authors.lastname: Gencic
authors.forename: David A
authors.initials: DA
authors.lastname: Grahame
authors.forename: Stephen P
authors.initials: SP
authors.lastname: Cramer
mesh_terms: D000445:Aldehyde Oxidoreductases / Q000737:chemistry* / Q000378:metabolism; D001665:Binding Sites; D002942:Circular Dichroism / Q000379:methods; D017020:Methanosarcina / Q000201:enzymology*; D009097:Multienzyme Complexes / Q000737:chemistry* / Q000378:metabolism; D009532:Nickel / Q000737:chemistry* / Q000378:metabolism; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013052:Spectrometry, X-Ray Emission / Q000379:methods
chemical_list: D009097:Multienzyme Complexes; D021122:Protein Subunits; D011994:Recombinant Proteins; D009532:Nickel; D000445:Aldehyde Oxidoreductases; C019823:carbon monoxide dehydrogenase