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1.
Chem Commun (Camb) ; 57(35): 4279-4282, 2021 Apr 29.
Article in English | MEDLINE | ID: mdl-33913973

ABSTRACT

We have fabricated Pt-Ca nanoparticles with oxygen reduction reaction catalytic activity via a sodium vapor-induced synthesis method. Prior addition of NaCl to form a eutectic mixture of CaCl2 and NaCl facilitated the formation of intermetallic Pt2Ca nanoparticles. Pt3Mg and Pt5Sr nanoparticles also were suggested to be producible.

2.
Article in English | MEDLINE | ID: mdl-20606269

ABSTRACT

Bilirubin oxidase (BOD), a multicopper oxidase found in Myrothecium verrucaria, catalyzes the oxidation of bilirubin to biliverdin. Oxygen is the electron acceptor and is reduced to water. BOD is used for diagnostic analysis of bilirubin in serum and has attracted considerable attention as an enzymatic catalyst for the cathode of biofuel cells that work under neutral conditions. Here, the crystal structure of BOD is reported for the first time. Blue bipyramid-shaped crystals of BOD obtained in 2-methyl-2,4-pentanediol (MPD) and ammonium sulfate solution were merohedrally twinned in space group P6(3). Structure determination was achieved by the single anomalous diffraction (SAD) method using the anomalous diffraction of Cu atoms and synchrotron radiation and twin refinement was performed in the resolution range 33-2.3 A. The overall organization of BOD is almost the same as that of other multicopper oxidases: the protein is folded into three domains and a total of four copper-binding sites are found in domains 1 and 3. Although the four copper-binding sites were almost identical to those of other multicopper oxidases, the hydrophilic Asn residue (at the same position as a hydrophobic residue such as Leu in other multicopper oxidases) very close to the type I copper might contribute to the characteristically high redox potential of BOD.


Subject(s)
Ascomycota/enzymology , Oxidoreductases Acting on CH-CH Group Donors/chemistry , Binding Sites , Copper/chemistry , Crystallography, X-Ray , Models, Molecular , Protein Structure, Tertiary
3.
Anal Chem ; 81(22): 9383-7, 2009 Nov 15.
Article in English | MEDLINE | ID: mdl-19908905

ABSTRACT

This paper describes a batch-type coulometric d-fructose biosensor based on direct electron transfer reaction of d-fructose dehydrogenase (FDH) adsorbed on a porous carbon electrode surface. The adsorbed-FDH electrodes catalyzed the electrochemical two-electron oxidation of d-fructose to 5-keto-d-fructose without a mediator. Nanostructured carbon particle-modified electrodes were used for the coulometric d-fructose biosensor to enhance the catalytic current density. The electric charge for the d-fructose oxidation gained by the biocoulometric measurement was in good agreement with the theoretical value corresponding to d-fructose amount in the range from 1 to 100 mM with a sample volume of 1 muL. This method is also applicable to the determination of several oligo/polysaccharides containing the d-fructose unit, in combination with specific hydrolases to yield d-fructose. An example was demonstrated by sucrose determination in which the electrode modified with FDH and invertase was used as a working electrode. To address the problem of electroactive interferences such as ascorbate, the electric charge at the FDH-free electrode was subtracted from the total charge obtained at the FDH-adsorbed electrode. The d-fructose concentrations in several beverages were successfully determined with this method.


Subject(s)
Chemistry Techniques, Analytical/methods , Electrochemical Techniques/methods , Fructose/analysis , Oxidoreductases/metabolism , Fructose/analogs & derivatives , Fructose/metabolism
4.
J Mol Biol ; 373(1): 141-52, 2007 Oct 12.
Article in English | MEDLINE | ID: mdl-17804014

ABSTRACT

CueO is a multicopper oxidase (MCO) that is involved in the homeostasis of Cu in Escherichia coli and is the sole cuprous oxidase to have ever been found. Differing from other MCOs, the substrate-binding site of CueO is deeply buried under a methionine-rich helical region including alpha-helices 5, 6, and 7 that interfere with the access of organic substrates. We deleted the region Pro357-His406 and replaced it with a Gly-Gly linker. The crystal structures of a truncated mutant in the presence and in the absence of excess Cu(II) indicated that the scaffold of the CueO molecule and metal-binding sites were reserved in comparison with those of CueO. In addition, the high thermostability of the protein molecule and its spectroscopic and magnetic properties due to four Cu centers were also conserved after truncation. As for functions, the cuprous oxidase activity of the mutant was reduced to ca 10% that of recombinant CueO owing to the decrease in the affinity of the labile Cu site for Cu(I) ions, although activities for laccase substrates such as 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid), p-phenylenediamine, and 2,6-dimethoxyphenol increased due to changes in the access of these organic substrates to the type I Cu site. The present engineering of CueO indicates that the methionine-rich alpha-helices function as a barrier to the access of bulky organic substrates, which provides CueO with specificity as a cuprous oxidase.


Subject(s)
Escherichia coli Proteins/chemistry , Escherichia coli Proteins/metabolism , Oxidoreductases/chemistry , Oxidoreductases/metabolism , Protein Structure, Tertiary , Binding Sites , Copper/metabolism , Crystallography, X-Ray , Enzyme Stability , Escherichia coli Proteins/genetics , Hydrogen-Ion Concentration , Models, Molecular , Molecular Sequence Data , Oxidoreductases/genetics , Protein Folding , Substrate Specificity , Temperature
5.
Phys Chem Chem Phys ; 9(15): 1793-801, 2007 Apr 21.
Article in English | MEDLINE | ID: mdl-17415490

ABSTRACT

One-compartment biofuel cells without separators have been constructed, in which d-fructose dehydrogenase (FDH) from Gluconobacter sp. and laccase from Trametes sp. (TsLAC) work as catalysts of direct electron transfer (DET)-type bioelectrocatalysis in the two-electron oxidation of d-fructose and four-electron reduction of dioxygen as fuels, respectively. FDH adsorbs strongly and stably on Ketjen black (KB) particles that have been modified on carbon papers (CP) and produces the catalytic current with the maximum density of about 4 mA cm(-2) without mediators at pH 5. The catalytic wave of the d-fructose oxidation is controlled by the enzyme kinetics. The location and the shape of the catalytic waves suggest strongly that the electron is directly transferred to the KB particles from the heme c site in FDH, of which the formal potential has been determined to be 39 mV vs. Ag|AgCl|sat. KCl. Electrochemistry of three kinds of multi-copper oxidases has also been investigated and TsLAC has been selected as the best one of the DET-type bioelectrocatalyst for the four-electron reduction of dioxygen in view of the thermodynamics and kinetics at pH 5. In the DET-type bioelectrocatalysis, the electron from electrodes seems to be transferred to the type I copper site of multi-copper oxidases. TsLAC adsorbed on carbon aerogel (CG) particles with an average pore size of 22 nm, that have been modified on CP electrodes, produces the catalytic reduction current of dioxygen with a density of about 4 mA cm(-2), which is governed by the mass transfer of the dissolved dioxygen. The FDH-adsorbed KB-modified CP electrodes and the TsLAC-adsorbed CG-modified CP electrodes have been combined to construct one-compartment biofuel cells without separators. The open-circuit voltage was 790 mV. The maximum current density of 2.8 mA cm(-2) and the maximum power density of 850 microW cm(-2) have been achieved at 410 mV of the cell voltage under stirring.


Subject(s)
Bioelectric Energy Sources , Carbohydrate Dehydrogenases/chemistry , Electrochemistry/instrumentation , Electrochemistry/methods , Fructose/chemistry , Gluconobacter/enzymology , Laccase/chemistry , Catalysis , Computer Simulation , Electron Transport , Equipment Design , Equipment Failure Analysis , Models, Chemical
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