127i Nuclear Magnetic Resonance Studies on the Interaction of Iodide Ion With Horseradish Peroxidase

Sakurada, Junji; Hosoya, Toshihiko; Shimizu, Tôru; Hatano, Masahiro

doi:10.1246/cl.1985.211

Cited by 9 publications

(7 citation statements)

References 11 publications

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“…As in the case of horseradish peroxidase (Sakurada et al, 1985), 1271 NMR studies demonstrated that an iodide ion binds to lactoperoxidase in two different manners: one, nonspecific and the other, specific (Figure 2). The former represents pH-independent binding and does not correlate with the specific activity of the enzyme, whereas the latter is controlled by the amino acid residue with pK, = 6.8 and correlates with the activity above pH 6.…”

Section: Discussionmentioning

confidence: 86%

“…As shown by the closed circles of Figure 2, deprotonation of an ionizing group with pKa = 6.8 f 0.1 (mean f SD) reduced the line-width broadening by the enzyme until a constant value (7.5 kHz) was obtained above pH 8.5. No pH effect on A I J~,~ of I-was observed in the absence of the enzyme (Sakurada et al, 1985). On the other hand, the pH effect on enzyme activity (triangles in Figure 2) revealed, on the basis of computer simulation, contribution of two amino acid residues with pKa = 4.8 f 0.2 and pKa = 7.1 f 0.1, the latter of which is very similar to the pK, = 6.8 mentioned above.…”

Section: Nmrmentioning

confidence: 84%

“…The binding of iodide to horseradish peroxidase was studied by spectrophotometric (Bjorksten, 1970), kinetic (Pommier et al, 1973), fluorometric (Ugarova et al, 1981), 12'1 NMR (Sakurada et al, 1985), and 'H NMR (Sakurada et al, 1987) techniques. The 'H NMR studies disclosed that 0006-2960/87/0426-6478$01 SO10 0 1987 American Chemical Society an iodide ion associates with the enzyme in the close vicinity of the heme peripheral 1-and 8-methyl groups at the place roughly in equal distance.…”

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confidence: 99%

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Proton and iodine-127 nuclear magnetic resonance studies on the binding of iodide by lactoperoxidase

Sakurada¹,

Takahashi²,

Shimizu³

et al. 1987

Biochemistry

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Interaction of an iodide ion with lactoperoxidase was studied by the use of 1H NMR, 127I NMR, and optical difference spectrum techniques. 1H NMR spectra demonstrated that a major broad hyperfine-shifted signal at about 60 ppm, which is ascribed to the heme peripheral methyl protons, was shifted toward high field by adding KI, indicating the binding of iodide to the active site of the enzyme; the dissociation constant was estimated to be 38 mM at pH 6.1. The binding was further detected by 127I NMR, showing no competition with cyanide. Both 1H NMR and 127I NMR revealed that the binding of iodide to the enzyme is facilitated by the protonation of an ionizable group with a pKa value of 6.0-6.8, which is presumably the distal histidyl residue. Optical difference spectra showed that the binding of an aromatic donor molecule to the enzyme is slightly but distinctly affected by adding KI. On the basis of these results, it was suggested that an iodide ion binds to lactoperoxidase outside the heme crevice but at the position close enough to interact with the distal histidyl residue which possibly mediates electron transport in the iodide oxidation reaction.

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Section: Discussionmentioning

confidence: 86%

Section: Nmrmentioning

confidence: 84%

mentioning

confidence: 99%

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Proton and iodine-127 nuclear magnetic resonance studies on the binding of iodide by lactoperoxidase

Sakurada¹,

Takahashi²,

Shimizu³

et al. 1987

Biochemistry

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“…Aromatic donors bind HRP in a hyrophobic pocket formed by haem peripheral 8-methyl, Ty-183 and Arg-185 residues [17], whereas iodide interacts near the haem moiety [5,12,18,19] at a site almost equidistant from haem peripheral 1-and 8-methyl groups [20]. Although the protonation of an ionizable group of PKa 4-4.6 appears to be necessary for iodide oxidation [11], Sakurada et al.…”

Section: Introductionmentioning

confidence: 99%

Mechanism of inhibition of horseradish peroxidase-catalysed iodide oxidation by EDTA

Bhattacharyya

Adak

Bandyopadhyay

et al. 1994

Biochemical Journal

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EDTA inhibits horseradish peroxidase (HRP)-catalysed iodide oxidation in a concentration and pH-dependent manner. It is more effective at pH 6 than at lower pH values. A plot of log Kiapp. values as a function of pH yields a sigmoidal curve from which a pKa value of 5.4 can be calculated for an ionizable group on the catalytically active HRP for EDTA inhibition. Among the structural analogues of EDTA, tetramethylethylenediamine (TEMED) is 80% as effective as EDTA, whereas the EDTA-Zn2+ chelate and EGTA are ineffective. Kinetic studies indicate that EDTA competitively inhibits iodide oxidation. Spectral studies show that EDTA can quickly reduce compound I to compound II, but reduction of preformed compound II to the native enzyme is relatively slow, as demonstrated by the time-dependent spectral shift from 417 nm to 402 nm through an isosbestic point at 408 nm. Under steady-state conditions, in a reaction mixture containing HRP, EDTA and H2O2, the enzyme remains in the compound-II form, with absorption maxima at 417, 527 and 556 nm. Direct evidence for one-electron oxidation of EDTA by HRP intermediates is provided by the appearance of an e.s.r. signal of a 5,5-dimethyl-1-pyrroline N-oxide (spin trap)-EDTA radical adduct [aN (hyperfine splitting constant) = 1.5 mT] in e.s.r. studies. The signal intensity, however, decreases in the presence of iodide. The KD of the HRP-EDTA complex obtained from optical difference spectroscopy increases with an increase in iodide concentration, and the double-reciprocal plot for EDTA binding indicates that EDTA and iodide compete for the same binding site for oxidation. We suggest that EDTA inhibits iodide oxidation by acting as an electron donor.

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“…Because it is very difficult to study the binding to compound I that causes the reaction, the binding of these ions to peroxidases in the resting state was examined by spectrophotometric (4,7,8), kinetic (9,10), fluorometric (11), 127 I NMR (12,13), 15 N NMR (14 -17), 13 C NMR (17), 1 H NMR (13)(14)(15)(16)(17)(18)(19), and optical difference spectroscopy (20 -22) techniques. The actual site of the binding of these ions to the enzymes and the mechanism of electron transfer from these ions to the heme irons have yet to be clarified.…”

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confidence: 99%

Binding of Iodide to Arthromyces ramosus Peroxidase Investigated with X-ray Crystallographic Analysis, 1H and 127I NMR Spectroscopy, and Steady-state Kinetics

Fukuyama

Sato

Itakura

et al. 1997

Journal of Biological Chemistry

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The site and characteristics of iodide binding to Arthromyces ramosus peroxidase were examined by x-ray crystallographic analysis, 1 H and 127 I NMR, and kinetic studies. X-ray analysis of an A. ramosus peroxidase crystal soaked in a KI solution at pH 5.5 showed that a single iodide ion is located at the entrance of the access channel to the distal side of the heme and lies between the two peptide segments, Phe

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127i Nuclear Magnetic Resonance Studies on the Interaction of Iodide Ion With Horseradish Peroxidase

Cited by 9 publications

References 11 publications

Proton and iodine-127 nuclear magnetic resonance studies on the binding of iodide by lactoperoxidase

Proton and iodine-127 nuclear magnetic resonance studies on the binding of iodide by lactoperoxidase

Mechanism of inhibition of horseradish peroxidase-catalysed iodide oxidation by EDTA

Binding of Iodide to Arthromyces ramosus Peroxidase Investigated with X-ray Crystallographic Analysis, 1H and 127I NMR Spectroscopy, and Steady-state Kinetics

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