The Experts below are selected from a list of 36927 Experts worldwide ranked by ideXlab platform

Jens Ulstrup - One of the best experts on this subject based on the ideXlab platform.

  • long range interfacial electrochemical electron transfer of pseudomonas aeruginosa azurin gold nanoparticle hybrid systems
    Journal of Physical Chemistry C, 2009
    Co-Authors: Palle Skovhus Jensen, Qijin Chi, Jingdong Zhang, Jens Ulstrup
    Abstract:

    We have prepared a “hybrid” of the blue Copper Protein azurin (Pseudomonas aeruginosa) and a 3 nm gold nanoparticle (AuNP). The AuNP/azurin hybrid was assembled on a Au(111)-electrode surface in a two-step process. The AuNP was first attached to the Au(111) electrode via Au−S chemisorption of a 4,4′-biphenyldithiol (4,4′-BPDT) monolayer. This was followed by 1-decanethiol modification of the bound AuNP and hydrophobic binding of azurin to the AuNP. The Au(111)/AuNP/azurin system was characterized by atomic force microscopy (AFM), cyclic voltammetry (CV), and in situ electrochemical scanning tunneling microscopy (in situ STM). AFM and STM point to the feasibility of preparing both dense and sparsely populated AuNP monolayers. CV shows two pairs of voltammetric peaks at high scan rates, both around the azurin equilibrium potential. One pair of redox peaks follows closely that of azurin hydrophobically immobilized directly on a Au(111)/1-tetradecanethiol reference surface. The other pair, tentatively assigne...

  • long range interfacial electron transfer of metalloProteins based on molecular wiring assemblies
    Faraday Discussions, 2006
    Co-Authors: Qijin Chi, Jingdong Zhang, Palle Skovhus Jensen, Hans Erik Molager Christensen, Jens Ulstrup
    Abstract:

    We address some physical features associated with long-range interfacial electron transfer (ET) of metalloProteins in both electrochemical and electrochemical scanning tunneling microscopy (ECSTM) configurations, which offer a brief foundation for understanding of the ET mechanisms. These features are illustrated experimentally by new developments of two systems with the blue Copper Protein azurin and enzyme nitrite reductase as model metalloProteins. Azurin and nitrite reductase were assembled on Au(111) surfaces by molecular wiring to establish effective electronic coupling between the redox centers in the Proteins and the electrode surface for ET and biological electrocatalysis. With such assemblies, interfacial ET proceeds through chemically defined and well oriented sites and parallels biological ET. In the case of azurin, the ET properties can be characterized comprehensively and even down to the single-molecule level with direct observation of redox-gated electron tunnelling resonance. Molecular wiring using a π-conjugated thiol is suitable for assembling monolayers of the enzyme with catalytic activity well-retained. The catalytic mechanism involves multiple-ET steps including both intramolecular and interfacial processes. Interestingly, ET appears to exhibit a substrate-gated pattern observed preliminarily in both voltammetry and ECSTM.

  • long range Protein electron transfer observed at the single molecule level in situ mapping of redox gated tunneling resonance
    Proceedings of the National Academy of Sciences of the United States of America, 2005
    Co-Authors: Qijin Chi, Ole Farver, Jens Ulstrup
    Abstract:

    A biomimetic long-range electron transfer (ET) system consisting of the blue Copper Protein azurin, a tunneling barrier bridge, and a gold single-crystal electrode was designed on the basis of molecular wiring self-assembly principles. This system is sufficiently stable and sensitive in a quasi-biological environment, suitable for detailed observations of long-range Protein interfacial ET at the nanoscale and single-molecule levels. Because azurin is located at clearly identifiable fixed sites in well controlled orientation, the ET configuration parallels biological ET. The ET is nonadiabatic, and the rate constants display tunneling features with distance-decay factors of 0.83 and 0.91 A–1 in H2O and D2O, respectively. Redox-gated tunneling resonance is observed in situ at the single-molecule level by using electrochemical scanning tunneling microscopy, exhibiting an asymmetric dependence on the redox potential. Maximum resonance appears around the equilibrium redox potential of azurin with an on/off current ratio of ≈9. Simulation analyses, based on a two-step interfacial ET model for the scanning tunneling microscopy redox process, were performed and provide quantitative information for rational understanding of the ET mechanism.

  • ordered assembly and controlled electron transfer of the blue Copper Protein azurin at gold 111 single crystal substrates
    Journal of Physical Chemistry B, 2001
    Co-Authors: Qijin Chi, Jingdong Zhang, Jens Enevold Thaulov Andersen, Jens Ulstrup
    Abstract:

    We have shown that Pseudomonas aeruginosa azurin can be immobilized on alkanethiol monolayers self-assembled on Au(111). Immobilization is achieved through hydrophobic interactions between the hydrophobic area around the Copper atom in azurin and methyl heads of alkanethiol to form submonolayers or monolayers. In this orientation mode azurin molecules on Au(111) are oriented with the redox center (Copper atom) facing the electrode surface. This is opposite to the orientation of azurin on bare gold which is via a surface disulfide group such as recently reported. Scanning tunneling microscopy (STM) with molecular resolution reveals that both well-ordered alkanethiol and Protein adlayers are present. Adsorbed azurin molecules exhibit high stability and retain electron transfer (ET) function. Long-range interfacial ET between azurin and Au(111) across variable-length alkanethiol bridges was systematically investigated by different electrochemical techniques. Distance-dependent ET can be controlled by adjusti...

  • molecular monolayers and interfacial electron transfer of pseudomonas aeruginosa azurin on au 111
    Journal of the American Chemical Society, 2000
    Co-Authors: Qijin Chi, Jingdong Zhang, Jens Enevold Thaulov Andersen, Esben Peter Friis, Gerard W Canters, Jens Ulrik Nielsen, Ib Chorkendorff, Jens Ulstrup
    Abstract:

    We provide a comprehensive approach to the formation and characterization of molecular monolayers of the blue Copper Protein Pseudomonas aeruginosa azurin on Au(111) in aqueous ammonium acetate solution. Main issues are adsorption patterns, reductive desorption, properties of the double layer, and long-range electrochemical electron transfer between the electrode and the Copper center. Voltammetry, electrochemical impedance spectroscopy (EIS), in situ scanning tunneling microscopy (STM), and X-ray photoelectron spectroscopy (XPS) have been employed to disclose features of these issues. Zn-substituted azurin, cystine, and 1-butanethiol are investigated for comparison. Cyclic voltammetric and capacitance measurements show qualitatively that azurin is adsorbed at submicromolar concentrations over a broad potential range. The characteristics of reductive desorption suggest that azurin is adsorbed via its disulfide group to form a monolayer. The adsorption of this Protein on Au(111) via a gold−sulfur binding m...

Qijin Chi - One of the best experts on this subject based on the ideXlab platform.

  • long range interfacial electrochemical electron transfer of pseudomonas aeruginosa azurin gold nanoparticle hybrid systems
    Journal of Physical Chemistry C, 2009
    Co-Authors: Palle Skovhus Jensen, Qijin Chi, Jingdong Zhang, Jens Ulstrup
    Abstract:

    We have prepared a “hybrid” of the blue Copper Protein azurin (Pseudomonas aeruginosa) and a 3 nm gold nanoparticle (AuNP). The AuNP/azurin hybrid was assembled on a Au(111)-electrode surface in a two-step process. The AuNP was first attached to the Au(111) electrode via Au−S chemisorption of a 4,4′-biphenyldithiol (4,4′-BPDT) monolayer. This was followed by 1-decanethiol modification of the bound AuNP and hydrophobic binding of azurin to the AuNP. The Au(111)/AuNP/azurin system was characterized by atomic force microscopy (AFM), cyclic voltammetry (CV), and in situ electrochemical scanning tunneling microscopy (in situ STM). AFM and STM point to the feasibility of preparing both dense and sparsely populated AuNP monolayers. CV shows two pairs of voltammetric peaks at high scan rates, both around the azurin equilibrium potential. One pair of redox peaks follows closely that of azurin hydrophobically immobilized directly on a Au(111)/1-tetradecanethiol reference surface. The other pair, tentatively assigne...

  • long range interfacial electron transfer of metalloProteins based on molecular wiring assemblies
    Faraday Discussions, 2006
    Co-Authors: Qijin Chi, Jingdong Zhang, Palle Skovhus Jensen, Hans Erik Molager Christensen, Jens Ulstrup
    Abstract:

    We address some physical features associated with long-range interfacial electron transfer (ET) of metalloProteins in both electrochemical and electrochemical scanning tunneling microscopy (ECSTM) configurations, which offer a brief foundation for understanding of the ET mechanisms. These features are illustrated experimentally by new developments of two systems with the blue Copper Protein azurin and enzyme nitrite reductase as model metalloProteins. Azurin and nitrite reductase were assembled on Au(111) surfaces by molecular wiring to establish effective electronic coupling between the redox centers in the Proteins and the electrode surface for ET and biological electrocatalysis. With such assemblies, interfacial ET proceeds through chemically defined and well oriented sites and parallels biological ET. In the case of azurin, the ET properties can be characterized comprehensively and even down to the single-molecule level with direct observation of redox-gated electron tunnelling resonance. Molecular wiring using a π-conjugated thiol is suitable for assembling monolayers of the enzyme with catalytic activity well-retained. The catalytic mechanism involves multiple-ET steps including both intramolecular and interfacial processes. Interestingly, ET appears to exhibit a substrate-gated pattern observed preliminarily in both voltammetry and ECSTM.

  • long range Protein electron transfer observed at the single molecule level in situ mapping of redox gated tunneling resonance
    Proceedings of the National Academy of Sciences of the United States of America, 2005
    Co-Authors: Qijin Chi, Ole Farver, Jens Ulstrup
    Abstract:

    A biomimetic long-range electron transfer (ET) system consisting of the blue Copper Protein azurin, a tunneling barrier bridge, and a gold single-crystal electrode was designed on the basis of molecular wiring self-assembly principles. This system is sufficiently stable and sensitive in a quasi-biological environment, suitable for detailed observations of long-range Protein interfacial ET at the nanoscale and single-molecule levels. Because azurin is located at clearly identifiable fixed sites in well controlled orientation, the ET configuration parallels biological ET. The ET is nonadiabatic, and the rate constants display tunneling features with distance-decay factors of 0.83 and 0.91 A–1 in H2O and D2O, respectively. Redox-gated tunneling resonance is observed in situ at the single-molecule level by using electrochemical scanning tunneling microscopy, exhibiting an asymmetric dependence on the redox potential. Maximum resonance appears around the equilibrium redox potential of azurin with an on/off current ratio of ≈9. Simulation analyses, based on a two-step interfacial ET model for the scanning tunneling microscopy redox process, were performed and provide quantitative information for rational understanding of the ET mechanism.

  • ordered assembly and controlled electron transfer of the blue Copper Protein azurin at gold 111 single crystal substrates
    Journal of Physical Chemistry B, 2001
    Co-Authors: Qijin Chi, Jingdong Zhang, Jens Enevold Thaulov Andersen, Jens Ulstrup
    Abstract:

    We have shown that Pseudomonas aeruginosa azurin can be immobilized on alkanethiol monolayers self-assembled on Au(111). Immobilization is achieved through hydrophobic interactions between the hydrophobic area around the Copper atom in azurin and methyl heads of alkanethiol to form submonolayers or monolayers. In this orientation mode azurin molecules on Au(111) are oriented with the redox center (Copper atom) facing the electrode surface. This is opposite to the orientation of azurin on bare gold which is via a surface disulfide group such as recently reported. Scanning tunneling microscopy (STM) with molecular resolution reveals that both well-ordered alkanethiol and Protein adlayers are present. Adsorbed azurin molecules exhibit high stability and retain electron transfer (ET) function. Long-range interfacial ET between azurin and Au(111) across variable-length alkanethiol bridges was systematically investigated by different electrochemical techniques. Distance-dependent ET can be controlled by adjusti...

  • molecular monolayers and interfacial electron transfer of pseudomonas aeruginosa azurin on au 111
    Journal of the American Chemical Society, 2000
    Co-Authors: Qijin Chi, Jingdong Zhang, Jens Enevold Thaulov Andersen, Esben Peter Friis, Gerard W Canters, Jens Ulrik Nielsen, Ib Chorkendorff, Jens Ulstrup
    Abstract:

    We provide a comprehensive approach to the formation and characterization of molecular monolayers of the blue Copper Protein Pseudomonas aeruginosa azurin on Au(111) in aqueous ammonium acetate solution. Main issues are adsorption patterns, reductive desorption, properties of the double layer, and long-range electrochemical electron transfer between the electrode and the Copper center. Voltammetry, electrochemical impedance spectroscopy (EIS), in situ scanning tunneling microscopy (STM), and X-ray photoelectron spectroscopy (XPS) have been employed to disclose features of these issues. Zn-substituted azurin, cystine, and 1-butanethiol are investigated for comparison. Cyclic voltammetric and capacitance measurements show qualitatively that azurin is adsorbed at submicromolar concentrations over a broad potential range. The characteristics of reductive desorption suggest that azurin is adsorbed via its disulfide group to form a monolayer. The adsorption of this Protein on Au(111) via a gold−sulfur binding m...

Jingdong Zhang - One of the best experts on this subject based on the ideXlab platform.

  • long range interfacial electrochemical electron transfer of pseudomonas aeruginosa azurin gold nanoparticle hybrid systems
    Journal of Physical Chemistry C, 2009
    Co-Authors: Palle Skovhus Jensen, Qijin Chi, Jingdong Zhang, Jens Ulstrup
    Abstract:

    We have prepared a “hybrid” of the blue Copper Protein azurin (Pseudomonas aeruginosa) and a 3 nm gold nanoparticle (AuNP). The AuNP/azurin hybrid was assembled on a Au(111)-electrode surface in a two-step process. The AuNP was first attached to the Au(111) electrode via Au−S chemisorption of a 4,4′-biphenyldithiol (4,4′-BPDT) monolayer. This was followed by 1-decanethiol modification of the bound AuNP and hydrophobic binding of azurin to the AuNP. The Au(111)/AuNP/azurin system was characterized by atomic force microscopy (AFM), cyclic voltammetry (CV), and in situ electrochemical scanning tunneling microscopy (in situ STM). AFM and STM point to the feasibility of preparing both dense and sparsely populated AuNP monolayers. CV shows two pairs of voltammetric peaks at high scan rates, both around the azurin equilibrium potential. One pair of redox peaks follows closely that of azurin hydrophobically immobilized directly on a Au(111)/1-tetradecanethiol reference surface. The other pair, tentatively assigne...

  • long range interfacial electron transfer of metalloProteins based on molecular wiring assemblies
    Faraday Discussions, 2006
    Co-Authors: Qijin Chi, Jingdong Zhang, Palle Skovhus Jensen, Hans Erik Molager Christensen, Jens Ulstrup
    Abstract:

    We address some physical features associated with long-range interfacial electron transfer (ET) of metalloProteins in both electrochemical and electrochemical scanning tunneling microscopy (ECSTM) configurations, which offer a brief foundation for understanding of the ET mechanisms. These features are illustrated experimentally by new developments of two systems with the blue Copper Protein azurin and enzyme nitrite reductase as model metalloProteins. Azurin and nitrite reductase were assembled on Au(111) surfaces by molecular wiring to establish effective electronic coupling between the redox centers in the Proteins and the electrode surface for ET and biological electrocatalysis. With such assemblies, interfacial ET proceeds through chemically defined and well oriented sites and parallels biological ET. In the case of azurin, the ET properties can be characterized comprehensively and even down to the single-molecule level with direct observation of redox-gated electron tunnelling resonance. Molecular wiring using a π-conjugated thiol is suitable for assembling monolayers of the enzyme with catalytic activity well-retained. The catalytic mechanism involves multiple-ET steps including both intramolecular and interfacial processes. Interestingly, ET appears to exhibit a substrate-gated pattern observed preliminarily in both voltammetry and ECSTM.

  • ordered assembly and controlled electron transfer of the blue Copper Protein azurin at gold 111 single crystal substrates
    Journal of Physical Chemistry B, 2001
    Co-Authors: Qijin Chi, Jingdong Zhang, Jens Enevold Thaulov Andersen, Jens Ulstrup
    Abstract:

    We have shown that Pseudomonas aeruginosa azurin can be immobilized on alkanethiol monolayers self-assembled on Au(111). Immobilization is achieved through hydrophobic interactions between the hydrophobic area around the Copper atom in azurin and methyl heads of alkanethiol to form submonolayers or monolayers. In this orientation mode azurin molecules on Au(111) are oriented with the redox center (Copper atom) facing the electrode surface. This is opposite to the orientation of azurin on bare gold which is via a surface disulfide group such as recently reported. Scanning tunneling microscopy (STM) with molecular resolution reveals that both well-ordered alkanethiol and Protein adlayers are present. Adsorbed azurin molecules exhibit high stability and retain electron transfer (ET) function. Long-range interfacial ET between azurin and Au(111) across variable-length alkanethiol bridges was systematically investigated by different electrochemical techniques. Distance-dependent ET can be controlled by adjusti...

  • molecular monolayers and interfacial electron transfer of pseudomonas aeruginosa azurin on au 111
    Journal of the American Chemical Society, 2000
    Co-Authors: Qijin Chi, Jingdong Zhang, Jens Enevold Thaulov Andersen, Esben Peter Friis, Gerard W Canters, Jens Ulrik Nielsen, Ib Chorkendorff, Jens Ulstrup
    Abstract:

    We provide a comprehensive approach to the formation and characterization of molecular monolayers of the blue Copper Protein Pseudomonas aeruginosa azurin on Au(111) in aqueous ammonium acetate solution. Main issues are adsorption patterns, reductive desorption, properties of the double layer, and long-range electrochemical electron transfer between the electrode and the Copper center. Voltammetry, electrochemical impedance spectroscopy (EIS), in situ scanning tunneling microscopy (STM), and X-ray photoelectron spectroscopy (XPS) have been employed to disclose features of these issues. Zn-substituted azurin, cystine, and 1-butanethiol are investigated for comparison. Cyclic voltammetric and capacitance measurements show qualitatively that azurin is adsorbed at submicromolar concentrations over a broad potential range. The characteristics of reductive desorption suggest that azurin is adsorbed via its disulfide group to form a monolayer. The adsorption of this Protein on Au(111) via a gold−sulfur binding m...

  • an approach to long range electron transfer mechanisms in metalloProteins in situ scanning tunneling microscopy with submolecular resolution
    Proceedings of the National Academy of Sciences of the United States of America, 1999
    Co-Authors: Esben Peter Friis, Jingdong Zhang, Jens Enevold Thaulov Andersen, Yu I Kharkats, A M Kuznetsov, Richard J Nichols, Jens Ulstrup
    Abstract:

    Abstract In situ scanning tunneling microscopy (STM) of redox molecules, in aqueous solution, shows interesting analogies and differences compared with interfacial electrochemical electron transfer (ET) and ET in homogeneous solution. This is because the redox level represents a deep indentation in the tunnel barrier, with possible temporary electronic population. Particular perspectives are that both the bias voltage and the overvoltage relative to a reference electrode can be controlled, reflected in spectroscopic features when the potential variation brings the redox level to cross the Fermi levels of the substrate and tip. The blue Copper Protein azurin adsorbs on gold(111) via a surface disulfide group. Well resolved in situ STM images show arrays of molecules on the triangular gold(111) terraces. This points to the feasibility of in situ STM of redox metalloProteins directly in their natural aqueous medium. Each structure also shows a central brighter contrast in the constant current mode, indicative of 2- to 4-fold current enhancement compared with the peripheral parts. This supports the notion of tunneling via the redox level of the Copper atom and of in situ STM as a new approach to long-range electron tunneling in metalloProteins.

Gerard W Canters - One of the best experts on this subject based on the ideXlab platform.

  • structural basis and mechanism of the inhibition of the type 3 Copper Protein tyrosinase from streptomyces antibioticus by halide ions
    Journal of Biological Chemistry, 2002
    Co-Authors: Armand W J W Tepper, Luigi Bubacco, Gerard W Canters
    Abstract:

    Abstract The inhibition of the type-3 Copper enzyme tyrosinase by halide ions was studied by kinetic and paramagnetic1H NMR methods. All halides are inhibitors in the conversion of l-3,4-dihydroxyphenylalanine (l-DOPA) with apparent inhibition constants that follow the order I− Cl− > Br− ≫ I−) and reduced (affinity I− > Br− > Cl− ≫ F−) enzyme. The paramagnetic 1H NMR of the oxidized enzyme complexed with the halides is consistent with a direct interaction of halide with the type-3 site and shows that the (Cu-His3)2coordination occurs in all halide-bound species. It is surmised that halides bridge both of the Copper ions in the active site. Fluoride and chloride are shown to bind only to the low pH form of oxidized tyrosinase, explaining the strong pH dependence of the inhibition by these ions. We further show that p-toluic acid and the bidentate transition state analogue, Kojic acid, displace chloride from the oxidized active site, whereas the monodentate substrate analogue,p-nitrophenol, forms a ternary complex with the enzyme and the chloride ion. On the basis of the experimental results, a model is formulated for the inhibitor action and for the reaction of diphenols with the oxidized enzyme.

  • the structural role of the Copper coordinating and surface exposed histidine residue in the blue Copper Protein azurin
    Journal of Molecular Biology, 2000
    Co-Authors: Lars J C Jeuken, Marcellus Ubbink, J H Bitter, Pieter Van Vliet, Wolfram Meyerklaucke, Gerard W Canters
    Abstract:

    Copper K-edge extended X-ray absorption fine structure (EXAFS) spectroscopy and 15N NMR relaxation studies were performed on samples of a variant azurin in which the surface-exposed histidine ligand of the Copper atom (His117) has been replaced by glycine. The experiments were performed to probe the structure of the active site and the Protein dynamics. The cavity in the Protein structure created by the His → Gly replacement could be filled by external ligands, which can either restore the spectroscopic properties of the original type-1 Copper site or create a new type-2 Copper site. The binding of external ligands occurs only when the Copper atom is in its oxidised state. In the reduced form, the binding is abolished. From the EXAFS experiments, it is concluded that for the oxidised type-1 Copper sites the Protein plus external ligand (L) provide an NSS∗L donor set deriving from His46, Cys112, Met121 and the external ligand. The type-2 Copper site features an S(N/O)3 donor set in which the S-donor derives from Cys112, one N-donor from His46 and the remaining two N or O donors from one or more external ligands. Upon reduction of the type-1 as well as the type-2 site, the external ligand drops out of the Copper site and the coordination reduces to 3-fold with an SS∗N donor set deriving from His46, Cys112 and Met121. The Cu-Sδ(Met) distance is reduced from about 3.2 to 2.3 A. Analysis of the NMR data shows that the hydrophobic patch around His117 has gained fluxionality when compared to wild-type azurin, which may explain why the His117Gly variant is able to accommodate a variety of external ligands of different sizes and with different chelating properties. On the other hand, the structure and dynamics of the β-sandwich, which comprises the main body of the Protein, is only slightly affected by the mutation. The unusually high reduction potential of the His117Gly azurin is discussed in light of the present results.

  • molecular monolayers and interfacial electron transfer of pseudomonas aeruginosa azurin on au 111
    Journal of the American Chemical Society, 2000
    Co-Authors: Qijin Chi, Jingdong Zhang, Jens Enevold Thaulov Andersen, Esben Peter Friis, Gerard W Canters, Jens Ulrik Nielsen, Ib Chorkendorff, Jens Ulstrup
    Abstract:

    We provide a comprehensive approach to the formation and characterization of molecular monolayers of the blue Copper Protein Pseudomonas aeruginosa azurin on Au(111) in aqueous ammonium acetate solution. Main issues are adsorption patterns, reductive desorption, properties of the double layer, and long-range electrochemical electron transfer between the electrode and the Copper center. Voltammetry, electrochemical impedance spectroscopy (EIS), in situ scanning tunneling microscopy (STM), and X-ray photoelectron spectroscopy (XPS) have been employed to disclose features of these issues. Zn-substituted azurin, cystine, and 1-butanethiol are investigated for comparison. Cyclic voltammetric and capacitance measurements show qualitatively that azurin is adsorbed at submicromolar concentrations over a broad potential range. The characteristics of reductive desorption suggest that azurin is adsorbed via its disulfide group to form a monolayer. The adsorption of this Protein on Au(111) via a gold−sulfur binding m...

  • in vivo studies disprove an obligatory role of azurin in denitrification in pseudomonas aeruginosa and show that azu expression is under control of rpos and anr
    Microbiology, 1997
    Co-Authors: Erik Vijgenboom, Julie E Busch, Gerard W Canters
    Abstract:

    Summary: The role of the blue Copper Protein azurin and cytochrome C551 as the possible electron donors to nitrite reductase in the dissimilatory nitrate reduction pathway in Pseudomonas aeruginosa have been investigated. It was shown by an in vivo approach with mutant strains of P. aeruginosa deficient in one or both of these electron-transfer Proteins that cytochrome C551, but not azurin, is functional in this pathway. Expression studies demonstrated the presence of azurin in both aerobic and anaerobic cultures. A sharp increase in azurin expression was observed when cultures were shifted from exponential to stationary phase. The stationary-phase sigma factor, σs, was shown to be responsible for this induction. In addition, one of the two promoters transcribing the azu gene was regulated by the anaerobic transcriptional regulator ANR. An azurin-deficient mutant was more sensitive to hydrogen peroxide and paraquat than the wild-type P. aeruginosa. These results suggest a physiological role of azurin in stress situations like those encountered in the transition to the stationary phase.

  • characterization and crystal structure of zinc azurin a by product of heterologous expression in escherichia coli of pseudomonas aeruginosa Copper azurin
    FEBS Journal, 1992
    Co-Authors: Herbert Nar, Mart Van De Kamp, Robert Huber, Albrecht Messerschmidt, Alexander Constantin Filippou, Manfred Barth, Michel Jaquinod, Gerard W Canters
    Abstract:

    Azurin*, a by-product of heterologous expression of the gene encoding the blue Copper Protein azurin from Pseudomonas aeruginosa in Escherichia coli, was characterized by chemical analysis and electrospray ionization mass spectrometry, and its structure determined by X-ray crystallography. It was shown that azurin* is native azurin with its Copper atom replaced by zinc in the metal binding site. Zinc is probably incorporated in the apo-Protein after its expression and transport into the periplasm. Holo-azurin can be reconstituted from azurin* by prolonged exposure of the Protein to high Copper ion concentrations or unfolding of the Protein and refolding in the presence of Copper ions. An X-ray crystallographic analysis of azurin* at 0.21-nm resolution revealed that the overall structure of azurin is not perturbed by the metal exchange. However, the geometry of the co-ordination sphere changes from trigonal bipyramidal in the case of Copper azurin to distorted tetrahedral for the zinc Protein. The Copper ligand Met121 is no longer co-ordinated to zinc which adopts a position close to the carbonyl oxygen atom from residue Gly45. The polypeptide structure surrounding the metal site undergoes moderate reorganization upon zinc binding. The largest displacement observed is for the carbonyl oxygen from residue Gly45, which is involved in Copper and zinc binding. It moves by 0.03 nm towards the zinc, thereby reducing its distance to the metal from 0.29 nm in the Copper Protein to 0.23 nm in the derivative.

Miguel A. De La Rosa - One of the best experts on this subject based on the ideXlab platform.

  • the cytochrome f plastocyanin complex as a model to study transient interactions between redox Proteins
    FEBS Letters, 2012
    Co-Authors: Isabel Cruzgallardo, Irene Diazmoreno, Antonio Diazquintana, Miguel A. De La Rosa
    Abstract:

    Transient complexes, with a lifetime ranging between microseconds and seconds, are essential for biochemical reactions requiring a fast turnover. That is the case of the interactions between Proteins engaged in electron transfer reactions, which are involved in relevant physiological processes such as respiration and photosynthesis. In the latter, the Copper Protein plastocyanin acts as a soluble carrier transferring electrons between the two membrane-embedded complexes cytochrome b6f and photosystem I. Here we review the combination of experimental efforts in the literature to unveil the functional and structural features of the complex between cytochrome f and plastocya- nin, which have widely been used as a suitable model for analyzing transient redox interactions. 2011 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  • the specificity in the interaction between cytochrome f and plastocyanin from the cyanobacterium nostoc sp pcc 7119 is mainly determined by the Copper Protein
    Biochemistry, 2007
    Co-Authors: Cristina Albarran, Miguel A. De La Rosa, Jose A Navarro, Manuel Hervas
    Abstract:

    : The plastocyanin-cytochrome f complex from Nostoc exhibits relevant structural differences when compared with the homologous complexes from other cyanobacteria and plants, with electrostatic and hydrophobic interactions being differently involved in each case. Here, five negatively charged residues of a recombinant form of cytochrome f from Nostoc have been replaced with either neutral or positively charged residues, and the effects of mutations on the kinetics of electron transfer to wild-type and mutant forms of plastocyanin have been measured by laser flash absorption spectroscopy. Cytochrome f mutants with some negative charges replaced with neutral residues exhibit an apparent electron transfer rate constant with wild-type plastocyanin similar to or slightly higher than that of the wild-type species, whereas the mutants with negative charges replaced with positive residues exhibit a significantly lower reactivity. Taken together, these results indicate that the effects of neutralizing residues at the electrostatically charged patch of cytochrome f are smaller than those previously observed for mutants of plastocyanin, thus suggesting that it is the Copper Protein which determines the specificity of the electrostatic interaction with the heme Protein. Moreover, cross reactions between mutants of both Proteins reveal the presence of some short-range specific electrostatic interactions. Our findings also make evident the fact that in Nostoc the main contribution to the electrostatic nature of the complex is provided by the small domain of cytochrome f.

  • a single arginyl residue in plastocyanin and in cytochrome c 6 from the cyanobacterium anabaena sp pcc 7119 is required for efficient reduction of photosystem i
    Journal of Biological Chemistry, 2001
    Co-Authors: Fernando P Molinaheredia, Manuel Hervas, Jose A Navarro, Miguel A. De La Rosa
    Abstract:

    Abstract Positively charged plastocyanin fromAnabaena sp. PCC 7119 was investigated by site-directed mutagenesis. The reactivity of its mutants toward photosystem I was analyzed by laser flash spectroscopy. Replacement of arginine at position 88, which is adjacent to the Copper ligand His-87, by glutamine and, in particular, by glutamate makes plastocyanin reduce its availability for transferring electrons to photosystem I. Such a residue in the Copper Protein thus appears to be isofunctional with Arg-64 (which is close to the heme group) in cytochrome c 6 from Anabaena(Molina-Heredia, F. P., Diaz-Quintana, A., Hervas, M., Navarro, J. A., and De la Rosa, M. A. (1999)J. Biol. Chem. 274, 33565–33570) andSynechocystis (De la Cerda, B., Diaz-Quintana, A., Navarro, J. A., Hervas, M., and De la Rosa, M. A. (1999) J. Biol. Chem. 274, 13292–13297). Other mutations concern specific residues of plastocyanin either at its positively charged east face (D49K, H57A, H57E, K58A, K58E, Y83A, and Y83F) or at its north hydrophobic pole (L12A, K33A, and K33E). Mutations altering the surface electrostatic potential distribution allow the Copper Protein to modulate its kinetic efficiency: the more positively charged the interaction site, the higher the rate constant. Whereas replacement of Tyr-83 by either alanine or phenylalanine has no effect on the kinetics of photosystem I reduction, Leu-12 and Lys-33 are essential for the reactivity of plastocyanin.