scholarly journals Dependence on freezing of the geometry and redox potential of type 1 and type 2 copper sites of Japanese-lacquer-tree (Rhus vernicifera) laccase

1981 ◽  
Vol 193 (2) ◽  
pp. 639-642 ◽  
Author(s):  
L Morpurgo ◽  
L Calabrese ◽  
A Desideri ◽  
G Rotilio
Keyword(s):  
Redox Potential ◽  
Hyperfine Splitting ◽  
Room Temperature ◽  
Redox Properties ◽  
Blue Copper ◽  
Hyperfine Splitting Constant ◽  
Splitting Constant ◽  
Rhus Vernicifera

The room-temperature e.p.r. spectrum of the Japanese-lacquer-tree (Rhus vernicifera) laccase shows A parallel (the hyperfine splitting constant) and g parallel values of both the Type 1 and Type 2 Cu appreciably different from those measured at liquid-N2 temperature. The geometry of the sites, as inferred from the room-temperature e.p.r. parameters, is more consistent with their redox properties. A rough correlation is found between A parallel and g parallel values and redox potential of the blue copper in several enzymes.

scholarly journals An e.p.r. study of the non-equivalence of the copper sites of caeruloplasmin

1986 ◽  
Vol 238 (1) ◽  
pp. 291-295 ◽  
Author(s):  
L Calabrese ◽  
M Carbonaro
Keyword(s):  
Slow Phase ◽  
Copper Binding ◽  
Blue Copper ◽  
Sequence Of Events ◽  
Copper Site ◽  
Hyperfine Splitting Constant ◽  
Type 1 Copper ◽  
Splitting Constant

The two Type 1 (blue) copper-binding sites of caeruloplasmin were spectroscopically differentiated by the kinetic analysis of the e.p.r. spectra during the redox cycle. One blue copper, with a hyperfine splitting constant (A parallel) of 6.8 mT, which was rapidly reduced, was not reoxidized by oxygen, whereas it was reoxidized by H2O2. The other blue copper (A parallel = 5.8 mT), which was reduced slowly, was rapidly reoxidized by either oxygen or H2O2. A conformational change of the Type 2 copper was concomitant with the fast reduction of Type 1 copper, whereas its reduction occurred during the slow phase. This sequence of events was reversed in the reoxidation step, that is, the Type 2 copper reappeared rapidly as the species with altered conformation and reverted to the symmetry typical of the native state in the slow phase. The specific reaction of a blue-copper site with the H2O2 can tentatively be related to the established ability of caeruloplasmin to prevent ‘oxidative’ attack of proteins and lipids.

scholarly journals Titrations with ferrocyanide of japanese-lacquer-tree (Rhus vernicifera) laccase and of the type 2 copper-depleted enzyme. Interrelation of the copper sites

1980 ◽  
Vol 187 (2) ◽  
pp. 367-370 ◽  
Author(s):  
L Morpurgo ◽  
M T Graziani ◽  
A Desideri ◽  
G Rotilio
Keyword(s):  
Absorption Band ◽  
Redox Potential ◽  
Electron Acceptor ◽  
Redox State ◽  
Redox Potentials ◽  
Rhus Vernicifera ◽  
Type 3 ◽  
Limiting Value

1. Redox titrations are reported of the metal centres in Japanese-lacquer-tree (Rhus vernicifera) laccase with ferrocyanide. 2. The redox potential of Type 1 Cu was found to increase with ferrocyanide concentration up to a limiting value similar to that for the Type 1 Cu in Type 2 Cu-depleted enzyme (which is independent of ferrocyanide concentration). 3. The redox potential of the two-electron acceptor (Type 3 Cu) is also independent of ferrocyanide concentration in Type 2 Cu-depleted enzyme and lower than values reported for the native enzyme. 4. The two-electron acceptor is present in the oxidized state in the Type 2 Cu-depleted enzyme, though the latter lacks the 330 nm absorption band. 5. The redox potential of Type 2 Cu also depends on ferrocyanide concentration, at least in the presence of azide. 6. The redox potentials are affected by freezing the solutions and/or addition of azide, the latter binding to Type 2 Cu with affinity dependent on the redox state of the two-electron acceptor.

Research on Effect of Alloy Elements on Equilibrium and Properties of Ni-Cr-Co Type Nickel-Based Superalloy

2016 ◽  
Vol 849 ◽  
pp. 513-519
Author(s):  
Qing Quan Zhang ◽  
Ming Yang Li ◽  
Ran Wei ◽  
Hui Yun Wu ◽  
Zhen Rui Li
Keyword(s):  
High Temperature ◽  
Room Temperature ◽  
High Temperature Strength ◽  
Nickel Based Superalloy ◽  
Type 3 ◽  
Room Temperature Strength ◽  
Super Alloy ◽  
Type Nickel

Ni-Cr-Co type Nickel-based super alloy Inconel 740H was studied. The effect of Nb, Al and Ti on the equilibrium of this alloy was analyzed by JMatPro software. The amount of Ti and Nb should be controlled by 1.50wt.%, and meanwhile, Al should be 1.0-2.0wt.%. If Mo and W were added the amount of Mo should be in the range of 1.0-2.0wt. %, and W should be about 1.0wt.%. Based on these results, three types of new alloys were designed, which contain Ni-Cr-Co-Mo type (1#), Ni-Cr-Co-W type (2#) and Ni-Cr-Co-Mo-W type (3#). Compared with the Ni-Cr-Co type Inconel 740H alloy, the room temperature strength, high temperature strength and high temperature durable performance of the three new alloys improved, which can provide the evidence and reference to optimize the chemical composition of Inconel 740H alloy, i.e., adding 1.50wt.% Mo and 1.0wt.% W individually or together.

scholarly journals Low-temperature resonance-Raman spectra of Japanese-lacquer-tree (Rhus vernicifera) laccase, type-2-copper-depleted laccase and H2O2-treated type-2-copper-depleted laccase

1983 ◽  
Vol 213 (2) ◽  
pp. 503-506 ◽  
Author(s):  
G Musci ◽  
A Desideri ◽  
L Morpurgo ◽  
A Garnier-Suillerot ◽  
L Tosi
Keyword(s):  
Raman Spectra ◽  
Strong Dependence ◽  
Resonance Raman ◽  
Local Symmetry ◽  
Structural Rearrangements ◽  
Blue Copper ◽  
Copper Site ◽  
Resonance Raman Spectra ◽  
Rhus Vernicifera

Resonance-Raman spectra of Japanese-lacquer-tree (Rhus vernicifera) laccase, type-2-copper-depleted laccase and the latter form treated with H2O2 were measured in liquid and frozen solution, on excitation into the 600 nm absorption band. Significant changes in intensity and/or frequency of the bands lying in the 370-430 cm-1 region were observed on freezing, indicating local structural rearrangements taking place at the blue copper site. These findings corroborate previous suggestions based on e.p.r. measurements and redox data [Morpurgo, Calabrese, Desideri & Rotilio (1981) Biochem. J. 193, 639-642]. They show the strong dependence of the physical properties of blue copper centres on local symmetry. Some conclusions on the origin of the Raman bands are also drawn.

scholarly journals The Blue Copper-Containing Nitrite Reductase fromAlcaligenes xylosoxidans: Cloning of the nirAGene and Characterization of the Recombinant Enzyme

1999 ◽  
Vol 181 (8) ◽  
pp. 2323-2329 ◽  
Author(s):  
Miguel Prudêncio ◽  
Robert R. Eady ◽  
Gary Sawers
Keyword(s):  
Amino Acid ◽  
Nitrite Reductase ◽  
Recombinant Enzyme ◽  
Leader Peptide ◽  
Resonance Spectroscopy ◽  
Gene Encoding ◽  
Blue Copper

ABSTRACT The nirA gene encoding the blue dissimilatory nitrite reductase from Alcaligenes xylosoxidans has been cloned and sequenced. To our knowledge, this is the first report of the characterization of a gene encoding a blue copper-containing nitrite reductase. The deduced amino acid sequence exhibits a high degree of similarity to other copper-containing nitrite reductases from various bacterial sources. The full-length protein included a 24-amino-acid leader peptide. The nirA gene was overexpressed inEscherichia coli and was shown to be exported to the periplasm. Purification was achieved in a single step, and analysis of the recombinant Nir enzyme revealed that cleavage of the signal peptide occurred at a position identical to that for the native enzyme isolated from A. xylosoxidans. The recombinant Nir isolated directly was blue and trimeric and, on the basis of electron paramagnetic resonance spectroscopy and metal analysis, possessed only type 1 copper centers. This type 2-depleted enzyme preparation also had a low nitrite reductase enzyme activity. Incubation of the periplasmic fraction with copper sulfate prior to purification resulted in the isolation of an enzyme with a full complement of type 1 and type 2 copper centers and a high specific activity. The kinetic properties of the recombinant enzyme were indistinguishable from those of the native nitrite reductase isolated from A. xylosoxidans. This rapid isolation procedure will greatly facilitate genetic and biochemical characterization of both wild-type and mutant derivatives of this protein.

scholarly journals Electron spin echo spectroscopic studies of type 1 and type 2 copper in Rhus vernicifera laccase and in Cucurbita pepo medullosa ascorbate oxidase

FEBS Letters ◽  
1981 ◽  
Vol 136 (1) ◽  
pp. 80-84 ◽  
Author(s):  
L. Avigliano ◽  
J.L. Davis ◽  
M.T. Graziani ◽  
A. Marchesini ◽  
W.B. Mims ◽  
...  
Keyword(s):  
Electron Spin ◽  
Cucurbita Pepo ◽  
Spin Echo ◽  
Electron Spin Echo ◽  
Spectroscopic Studies ◽  
Ascorbate Oxidase ◽  
Rhus Vernicifera

scholarly journals What properties of non-electrolyte solutions does a DNA molecule feel?

2021 ◽  
Vol 13 (2) ◽  
pp. 129-136
Author(s):  
George G. Malenkov ◽  
Keyword(s):  
Relative Humidity ◽  
Ethylene Glycol ◽  
Water Activity ◽  
Hyperfine Splitting ◽  
Solvent Polarity ◽  
Electrolyte Solutions ◽  
Water Solutions ◽  
Hyperfine Splitting Constant ◽  
Splitting Constant ◽  
In The Beginning

As early as in 1953 it was reported that at low relative humidity of the atmosphere (about 70 %) DNA films were in A-form, while at higher humidity (more than 80%) they were in B-form. Relative humidity of the atmosphere corresponds to the water activity in the system. It was found in the beginning of the 70th that DNA transfers to the A-form when non-electrolytes, such as monoatomic alcohols, dioxane, tetrahydrofurane were added to water. It was reasonable to suppose that B to A transition of DNA occurs in the non-electrolyte solutions at the same values af water activity as in the moist atmosphere. This prediction was borne out. But in water solutions of very polar non-electrolytes such as methanol and ethylene glycol B to A DNA does not occur even at very low water activity values. It was supposed that A form of DNA can arise only in the medium with sufficiently low polarity. We used hyperfine splitting constant (A) of a nitroxide spin label determined from the EPR spectra as a measure of the liquid solvent polarity. It was found that DNA transition into B form occurs when A reached a certain value. Polarity of methanol and ethylene glycol is much higher than that of the solutions in which B to A DNA transition takes place. This transition also occurs in water solutions of trifluoroethanol. But in these solutions the transition takes place at hyperfine splitting values, which are much greater than in other non-electrolyte solutions. Water activity in the zone of B-A DNA transition in trifluoroethanol solutions does not correspond water activity which was calculated for other nob-electrolyte solutions in which B-A transition is observed.

First Synthesis of C-Phenyl N-tert-Butyl [15N]nitrone (PBN-15N) for the EPR Spin Trapping Methodology

1996 ◽  
Vol 51 (1) ◽  
pp. 139-143 ◽  
Author(s):  
Yong-Kang Zhang
Keyword(s):  
Hyperfine Splitting ◽  
Spin Trap ◽  
Spin Trapping ◽  
Paramagnetic Resonance ◽  
Tert Butyl ◽  
Hyperfine Splitting Constant ◽  
Percent Increase ◽  
Splitting Constant ◽  
Epr Spin Trapping ◽  
Electron Paramagnetic

As compared to normal PBN, about fifty percent increase of electron paramagnetic resonance (EPR) spin trapping sensitivity has been gained by using a new 100% 15N-enriched spin trap, C-phenyl N-tert-butyl[15N]nitrone (PBN-15N). PBN-15N has been prepared by a convenient four-step route using ammonium-15N chloride as the starting material. This synthetic method produces 2-methyl-2-[15N]nitropropane which is useful for the synthesis of many other PBN-15N type spin traps for the purpose of increasing spin trapping sensitivity. EPR spin trapping with PBN-15N in benzene and in phosphate buffer has been investigated. The 15N hyperfine splitting constant (15N-hfsc) is larger than 14N-hfsc by 40%. The larger 15N-hfsc gives more opportunity to identify different radical addends within the same system.

scholarly journals Azide-binding studies reveal type 3 copper heterogeneity in ascorbate oxidase from the green zucchini squash (Cucurbita pepo)

1988 ◽  
Vol 251 (2) ◽  
pp. 441-446 ◽  
Author(s):  
L Casella ◽  
M Gullotti ◽  
G Pallanza ◽  
A Pintar ◽  
A Marchesini
Keyword(s):  
Cucurbita Pepo ◽  
Ascorbate Oxidase ◽  
Binding Studies ◽  
High Affinity ◽  
Hyperfine Splitting Constant ◽  
Splitting Constant ◽  
A Minor ◽  
Type 3 ◽  
Azide Binding

Titration of native ascorbate oxidase from green zucchini squash (Cucurbita pepo) with azide in 0.1 M-phosphate buffer, pH 6.8, exhibits a biphasic spectral behaviour. Binding of the anion with ‘high affinity’ (K greater than 5000 M-1) produces a broad increase of absorption in the 400-500 nm region (delta epsilon approximately 1000 M-1.cm-1) and c.d. activity in the 300-450 nm region, whereas azide binding with ‘low affinity’ (K approximately 100 M-1) is characterized by an intense absorption band at 420 nm (delta epsilon = 6000 M-1.cm-1), corresponding to negative c.d. activity and a decrease of absorption at 330 nm (delta epsilon = -2000 M-1.cm-1). The high-affinity binding involves a minor fraction of the protein containing Type 3 copper in the reduced state, and the spectral features of this azide adduct can be eliminated by treatment of the native enzyme with small amounts of H2O2, followed by dialysis before azide addition. As shown by e.s.r. spectroscopy, Type 2 copper is involved in both types of binding, its signal being converted into that of a species with small hyperfine splitting constant [12 mT (approximately 120 G)] in the case of the low-affinity azide adduct. The spectral similarities of the two types of azide adducts with the corresponding adducts formed by native laccase, which also exhibits Type 3 copper heterogeneity, are discussed.

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