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Komorowski, Lars; Anemüller, Stefan; Schäfer, Günter

doi:10.1023/a:1005668522801

Cited by 14 publications

(13 citation statements)

References 17 publications

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“…acidocaldarius . 1331 The UV–vis, EPR, and EXAFS spectroscopic characterizations as well as the reduction potentials measurments for these soluble truncates are consistent with each other (Table 13). 742,1339,1358,1361 To date, only the truncate from T. thermophilus has been successfully crystallized.…”

Section: Copper Redox Centers In Electron Transfer Processessupporting

confidence: 70%

“…acidocaldarius (SoxH), 1331 and a nitric oxide reductase (qCu A NOR) 1332,1333 to date (Figure 60). Interestingly, all of these proteins are terminal electron acceptors of ET processes; e.g., C c O is the terminal electron acceptor in aerobic respiration, and N 2 OR is the terminal electron acceptor in anaerobic respiration.…”

Section: Copper Redox Centers In Electron Transfer Processesmentioning

confidence: 99%

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Metalloproteins Containing Cytochrome, Iron–Sulfur, or Copper Redox Centers

et al. 2014

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Blue Type I Copper Centers vs Purple Cu A Centers 4439 5. Enzymes Employing a Combination of Different Types of Electron Transfer Centers 4439 5.1. Enzymes Using Both Heme and Cu as Electron Transfer Centers 4439 5.1.1. Cytochrome c and Cu A as Redox Partners to Cytochrome c Oxidases 4439 5.1.2. Cu A and Heme b as Redox Partners to Nitric Oxide Reductases 4440 5.1.3. Cytochrome c and Cu A as Redox Partners to Nitrous Oxide Reductases 4440 5.2. Enzymes Using Both Heme and Iron−Sulfur Clusters as Electron Transfer Centers 4440 5.2.1. As Redox Partners to the Cytochrome bc 1 Complex 4440 5.2.2. As Redox Partners to the Cytochrome b 6 f Complex 4442 5.2.3. As Redox Centers in Formate Dehydrogenases 4443 5.2.4. As Redox Centers in Nitrate Reductase 4443 6. Summary and Outlook 4443 Author Information 4445 Corresponding Author 4445 Author Contributions 4445 Notes 4445 Biographies 4445 Acknowledgments 4447 Abbreviations 4447 References 4447

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Section: Copper Redox Centers In Electron Transfer Processessupporting

confidence: 70%

Section: Copper Redox Centers In Electron Transfer Processesmentioning

confidence: 99%

Metalloproteins Containing Cytochrome, Iron–Sulfur, or Copper Redox Centers

et al. 2014

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Section: Introductionmentioning

confidence: 99%

“…16, 17, 19, 20 One strategy that has been employed to study Cu A sites involves expressing the soluble Cu A domain from C c O and reconstituting the Cu A site in vitro. 16, 25–27, 32–36 In a second strategy the Cu A ligands have been engineered in another cupredoxin protein followed by in vitro reconstitution of the binuclear site. 37–40 By replacing the Cu binding loop from T1Cu proteins with the Cu A loop from C c O, the Cu A site was successfully engineered into CyoA domain of a copper-less quinol oxidase (called Cu A -CyoA) and blue T1Cu proteins amicyanin (called Cu A Ami) and azurin (Az, called Cu A Az).…”

Section: Introductionmentioning

confidence: 99%

Binuclear Cu_A Formation in Biosynthetic Models of Cu_A in Azurin Proceeds via a Novel Cu(Cys)₂His Mononuclear Copper Intermediate

et al. 2015

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CuA is a binuclear electron transfer (ET) center found in cytochrome c oxidases (CcOs), nitrous oxide reductases (N2ORs), and nitric oxide reductase (NOR). In these proteins, the CuA centers facilitate efficient ET (kET > 104 s−1) under low thermodynamic driving forces (10–90 mV). While the structure and functional properties of CuA are well understood, a detailed mechanism of copper incorporation into the protein and the identity of the intermediates formed during the CuA maturation process are still lacking. Previous studies of the CuA assembly mechanism in vitro using a biosynthetic model CuA center in azurin (CuAAz) identified a novel intermediate X (Ix) during reconstitution of the binuclear site. However, due to the instability of Ix and the coexistence of other Cu centers, such as CuA’ and type 1 copper centers, the identity of this intermediate could not be established. Here, we report the mechanism of CuA assembly using variants of Glu114XCuAAz (X=Gly, Ala, Leu, Gln), the backbone carbonyl of which acts as a ligand to the CuA site, with a major focus on characterization of the novel intermediate Ix. We show that CuA assembly in these variants proceeds through several types of Cu centers, such as mononuclear red type 2 Cu, the novel intermediate Ix, and blue type 1 Cu. Our results show that the backbone flexibility of the Glu114 residue is an important factor in determining the rates of T2Cu→Ix formation, suggesting that the CuA formation is facilitated by swinging of the ligand loop, which internalizes the T2Cu capture complex to the protein interior. The kinetic data further suggests that the nature of the Glu114 side chain influences the timescales on which these intermediates are formed, the wavelengths of the absorption peaks, and how cleanly one intermediate is converted to another. Through careful understanding of these mechanism and optimization of the conditions, we have obtained Ix in ~80–85% population in these variants, which allowed us to employ UV-Vis, EPR, and EXAFS spectroscopic techniques to identify the Ix as a mononuclear Cu(Cys)2(His) complex. Since some of the intermediates have been proposed to be involved in the assembly of native CuA, these results shed light on the structural features of the important intermediates and mechanism of CuA formation.

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“…4,5 Additionally, Cu A is a cofactor in the terminal oxidase (SoxH) from Sulfolobus acidocaldarius, thermoacidophilic bacteria found in sulfur-rich acid springs. 6 The Cu A resting state is charge-delocalized featuring a [Cu 1.5 Cu 1.5 ] + core that is also the rst metalloenzyme observed with a very close metal-metal separation (2.44 Å). 7 The binuclear nature of the Cu A site is enforced by two bridging cysteine residues that link two His-Cu + units to produce a rigid Cu 2 S 2 diamond core (Cu-S distances z 2.2-2.4 Å) that enables fast electron transfer between the [Cu 1.5 Cu 1.5 ] + and [Cu I Cu I ] states.…”

Section: Introductionmentioning

confidence: 99%

Three coordinate models for the binuclear CuA electron-transfer site

Zhang

Warren

2013

Chem. Sci.

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Truly three coordinate Cu A synthetic models were developed by employing bulky N-heterocyclic carbene ligands along with simple C 3 and C 4 dithiolate linkers to emulate the role of histidine and cysteine ligands at trigonal copper sites. The reduced [Cu I Cu I ] models [NHC]Cu-S(CH 2 ) n S-Cu[NHC] (n ¼ 3 (3), 4 (4)) were characterized by NMR and X-ray crystallography. The structure of the reduced model with the C 3 linker closely mimics the low coordinate, trigonal environment at the reduced dinuclear Cu A site while the C 4 linker resulted in an "open" solid state structure with two coordinate Cu centers. Oxidized species 3 + and 4 + were observed by UV-vis and EPR spectroscopy upon the addition of [Cp 2 Fe] + to 3 and 4, generating spectroscopic features similar to those of the dinuclear [Cu 1.5 Cu 1.5 ] + Cu A resting state. Key spectroscopic features of 3 + and 4 + are low energy intervalence charge transfer bands centered near 1150 nm and EPR spectra indicating delocalization of the unpaired e À among two Cu centers.Electrochemistry of the oxidized [Cu 2 S 2 ] + models reveals reduction potentials (E 1/2 ¼ 0.11 V (3 + ); 0.36 V (4 + ) vs. NHE) similar to the Cu A site in CcO (E 1/2 ¼ 0.22-0.30 V vs. NHE).

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Cited by 14 publications

References 17 publications

Metalloproteins Containing Cytochrome, Iron–Sulfur, or Copper Redox Centers

Metalloproteins Containing Cytochrome, Iron–Sulfur, or Copper Redox Centers

Binuclear Cu_A Formation in Biosynthetic Models of Cu_A in Azurin Proceeds via a Novel Cu(Cys)₂His Mononuclear Copper Intermediate

Three coordinate models for the binuclear CuA electron-transfer site

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Untitled

Cited by 14 publications

References 17 publications

Metalloproteins Containing Cytochrome, Iron–Sulfur, or Copper Redox Centers

Metalloproteins Containing Cytochrome, Iron–Sulfur, or Copper Redox Centers

Binuclear CuA Formation in Biosynthetic Models of CuA in Azurin Proceeds via a Novel Cu(Cys)2His Mononuclear Copper Intermediate

Three coordinate models for the binuclear CuA electron-transfer site

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Binuclear Cu_A Formation in Biosynthetic Models of Cu_A in Azurin Proceeds via a Novel Cu(Cys)₂His Mononuclear Copper Intermediate