A comparative reactivity study of microperoxidases based on hemin, mesohemin and deuterohemin

Ryabova, Ekaterina S.; Rydberg, Patrik; Kolberg, Matthias; Harbitz, Espen; Barra, Anne Laure; Ryde, Ulf; Andersson, K. Kristoffer; Nordlander, Ebbe

doi:10.1016/j.jinorgbio.2004.12.020

Cited by 17 publications

(13 citation statements)

References 71 publications

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“…In order to quantify the peroxidase activity of salen cyclodextrin conjugates, ABTS was used as the substrate [56][57][58]. The reaction of ABTS with H 2 O 2 in the presence of 4a or 4b generates ABTS Å+ :…”

Section: Peroxidase Activitymentioning

confidence: 99%

New conjugates of superoxide dismutase/catalase mimetics with cyclodestrins

Lanza

Vecchio

2009

Journal of Inorganic Biochemistry

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Section: Peroxidase Activitymentioning

confidence: 99%

New conjugates of superoxide dismutase/catalase mimetics with cyclodestrins

Lanza

Vecchio

2009

Journal of Inorganic Biochemistry

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“…[23][24][25] Increased electron density on the metal center should increase the rate of heterolysis to Cpd I by enhancing the "push effect," 26 while also stabilizing the resulting Cpd I species. With peroxidases, meso-heme substitution leads to increases in peroxide cleavage.…”

Section: Peroxide Shunt Reactionsmentioning

confidence: 99%

The Second Step of the Nitric Oxide Synthase Reaction: Evidence for Ferric-Peroxo as the Active Oxidant

et al. 2008

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Nitric oxide synthase (NOS) is a P450 mono-oxygenase that catalyzes the oxidation of l-arginine to citrulline and NO through the stable intermediate N(G)-hydroxy-l-arginine (NHA). The oxidation of NHA by NOS is unique. There is little direct evidence in support of the nature of the heme bound oxidant [i.e., ferric-peroxo vs Fe(IV)O(por(*+))] responsible for this transformation. Previous work characterizing the H(2)O(2)-driven oxidation of NHA by NOS showed the formation of citrulline and the side product N(delta)-cyanoornithine (CN-orn). This led to the proposed involvement of a ferric-peroxo intermediate in the oxidation of NHA to citrulline. To test this hypothesis we used this model reaction to study the effects of pH, heme substitution, active site mutagenesis, and a fluorinated substrate analogue on the product distribution. Further, the oxidation of 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS) by H(2)O(2) and iNOS(heme) was used to probe the protein-catalyzed breakdown of peroxide to the Fe(IV)O(por(*+)) intermediate. At pH 6.5, 7.5, and 8.5 the peroxide shunt reaction forms 26 +/- 2, 36 +/- 1, and 51 +/- 1% citrulline, respectively. The rate of peroxidase activity, however, was negatively correlated to pH, with a peroxide breakdown rate of 13.1 +/- 0.3, 8.3 +/- 0.2, and 4.2 +/- 0.1 M(-1) s(-1) at pH 6.5, 7.5, and 8.5, respectively. Mutation of active site valine 346 to an alanine shifted the product distribution to 5.2 +/- 0.5% citrulline while enhancing the peroxide cleavage rate to 14.3 +/- 0.7 M(-1) s(-1). Substitution of the heme cofactor with iron mesoporphyrin IX (Fe-MPIX) alters the product distribution from 36 +/- 1% citrulline to 22 +/- 3% citrulline. Metal substitution with Mn results in the formation of 64.7 +/- 0.8% citrulline. Conversely, the electrophilic 4,4-difluoro-N(G)-hydroxy-l-arginine substrate analogue shifted the product distribution to 68.6 +/- 0.6% 4,4-difluorocitrulline. The peroxidase data provide insight into the chemical features of NOS that control the processing of the ferric-peroxo species to the Fe(IV)O(por(*+)) intermediate and help interpret the product distributions observed for the peroxide shunt under various conditions. In all cases, the ability of the protein to break down peroxide is negatively correlated with the formation of citrulline by the peroxide shunt. These results support the high valent Fe(IV)O(por(*+)) intermediate as the species responsible for CN-orn formation and are consistent with the involvement of the ferric-peroxo intermediate in the oxidation of NHA to citrulline.

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“…The ABTS proved to be particularly suitable as a substrate and it is commonly applied for the quantification of the enzymatic activity of peroxidases [33][34][35], because it serves to trap any hypervalent manganese-salen species as well as hydroxyl radical products. The reduced form of ABTS is colourless; by contrast, when ABTS is oxidised to the radical cation several characteristic absorption bands appear at 415, 650, 735 and 815 nm and the resulting solution is green [33,36].…”

Section: Peroxidase Assaysmentioning

confidence: 99%