pH Dependence of the oxidation of iodide by compound I of horseradish peroxidase

Biochemistry ◽  
1972 ◽  
Vol 11 (11) ◽  
pp. 2076-2082 ◽  
Author(s):  
R. Roman ◽  
H. B. Dunford
Keyword(s):  
Horseradish Peroxidase ◽  
Ph Dependence ◽  
Compound I

Kinetics of the oxidation of reduced nicotinamide adenine dinucleotide by horseradish peroxidase compounds I and II

1986 ◽  
Vol 64 (4) ◽  
pp. 323-327 ◽  
Author(s):  
Mohammed A. Kashem ◽  
H. Brian Dunford
Keyword(s):  
Horseradish Peroxidase ◽  
Active Site ◽  
Nicotinamide Adenine Dinucleotide ◽  
Ph Dependence ◽  
Transient State ◽  
Nicotinamide Adenine ◽  
Compound I ◽  
Single Ionization ◽  
Reduced Nicotinamide Adenine Dinucleotide ◽  
Kinetics Of

The transient state kinetics of the oxidation of reduced nicotinamide adenine dinucleotide (NADH) by horseradish peroxidase compound I and II (HRP-I and HRP-II) was investigated as a function of pH at 25.0 °C in aqueous solutions of ionic strength 0.11 using both a stopped-flow apparatus and a conventional spectrophotometer. In agreement with studies using many other substrates, the pH dependence of the HRP-I–NADH reaction can be explained in terms of a single ionization of pKa = 4.7 ± 0.5 at the active site of HRP-I. Contrary to studies with other substrates, the pH dependence of the HRP-H–NADH reaction can be interpreted in terms of a single ionization with pKa of 4.2 ± 1.4 at the active site of HRP-II. An apparent reversibility of the HRP-II–NADH reaction was observed. Over the pH range of 4–10 the rate constant for the reaction of HRP-I with NADH varied from 2.6 × 105 to5.6 × 102 M−1 s−1 and of HRP-II with NADH varied from 4.4 × 104 to 4.1 M−1 s−1. These rate constants must be taken into consideration to explain quantitatively the oxidase reaction of horseradish peroxidase with NADH.

Studies on Horseradish Peroxidase. XII. A Kinetic Study of the Oxidation of Sulfite and Nitrite by Compounds I and II

1973 ◽  
Vol 51 (4) ◽  
pp. 588-596 ◽  
Author(s):  
R. Roman ◽  
H. B. Dunford
Keyword(s):  
Horseradish Peroxidase ◽  
Ground State ◽  
Ph Dependence ◽  
Intermediate Formation ◽  
Ion Concentration ◽  
Compound I ◽  
Hydrogen Ion Concentration ◽  
Ph Range ◽  
Compound Ii ◽  
Kinetics Of

The kinetics of the oxidation of sulfite and nitrite by horseradish peroxidase compounds I and II have been studied as a function of pH at 25° and ionic strength 0.11. The pH dependence of the rate of the reaction between compound I and sulfite over the pH range 2–7 is interpreted in terms of two ground state enzyme dissociations with pka values of 5.1 and 3.3, and that for the compound II reaction with sulfite in terms of a single ground state enzyme dissociation with a pKa value of 3.9. Whereas the reaction between compound I and sulfite produces the native enzyme without the intermediate formation of compound II, the reaction of compound I with nitrite yields compound II. The second-order rate constants for the reactions of compounds I and II with nitrite increase linearly with increasing hydrogen ion concentration over the pH range 6–8.

Studies on Horseradish Peroxidase. XI. On the Nature of Compounds I and II as Determined from the Kinetics of the Oxidation of Ferrocyanide

1973 ◽  
Vol 51 (4) ◽  
pp. 582-587 ◽  
Author(s):  
M. L. Cotton ◽  
H. B. Dunford
Keyword(s):  
Horseradish Peroxidase ◽  
Rate Constant ◽  
Active Sites ◽  
Ph Dependence ◽  
Order Rate Constant ◽  
Second Order ◽  
Compound I ◽  
Order Rate ◽  
Compound Ii ◽  
Kinetics Of

In order to investigate the nature of compounds I and II of horseradish peroxidase, the kinetics were studied of ferrocyanide oxidation catalyzed by these compounds which were prepared from three different oxidizing agents. The pH dependence of the apparent second-order rate constant for ferrocyanide oxidation by compound I, prepared from ethyl hydroperoxide and m-chloroperbenzoic acid, was interpreted in terms of an ionization on the enzyme with a pKa = 5.3, identical to that reported previously for hydrogen peroxide. The second-order rate constant for the compound II-ferrocyanide reaction also showed the same pH dependence for the three oxidizing substrates. However, with more accurate results, the compound II-ferrocyanide reaction was reinterpreted in terms of a single ionization with pKa = 8.5. The same dependence of ferrocyanide oxidation on pH suggests structurally identical active sites for compounds I and II prepared from the three different oxidizing substrates.

Horseradish peroxidase. XXXII. pH dependence of the oxidation of L-(−)-tyrosine by compound I

1978 ◽  
Vol 56 (12) ◽  
pp. 1115-1119 ◽  
Author(s):  
I. Ralston ◽  
H. B. Dunford
Keyword(s):  
Horseradish Peroxidase ◽  
Ph Dependence ◽  
Nonlinear Least Squares ◽  
Compound I ◽  
Ph Profile ◽  
Rate Versus ◽  
Tyrosine Oxidation ◽  
Phenolic Group ◽  
Ph Range ◽  
Single Enzyme

The rate of oxidation of L-(−)-tyrosine by horseradish peroxidase compound I has been studied as a function of pH at 25 °C and ionic strength 0.11. Over the pH range of 3.20–11.23 major effects of three ionizations were observed. The pKa values of the phenolic (pKa = 10.10) and amino (pKa = 9.21) dissociations of tyrosine and a single enzyme ionization (pKa = 5.42) were determined from nonlinear least squares analysis of the log rate versus pH profile. It was noted that the less acidic form of the enzyme was most reactive; hence, the reaction is described as base catalyzed. The rate of tyrosine oxidation falls rapidly with the deprotonation of the phenolic group.

scholarly journals Iodide modulation of the EDTA-induced iodine reductase activity of horseradish peroxidase by interaction at or near the EDTA-binding site

1993 ◽  
Vol 289 (2) ◽  
pp. 575-580 ◽  
Author(s):  
D K Bhattacharyya ◽  
U Bandyopadhyay ◽  
R Chatterjee ◽  
R K Banerjee
Keyword(s):  
Horseradish Peroxidase ◽  
Binding Site ◽  
Reductase Activity ◽  
Electron Flow ◽  
Ph Dependence ◽  
Compound I ◽  
Native Form ◽  
Iodide Oxidation ◽  
Equilibrium Dissociation ◽  
Haem Iron

Horseradish peroxidase (HRP) catalyses the reduction of iodinium ion (I+) to iodide by H2O2 in the presence of EDTA. I+ reduction occurs optimally at pH 6 whereas the enzyme catalyses iodide oxidation optimally at pH 3.5. Thus the two activities reside on the same enzyme with two characteristic pH optima. Iodide modulates the expression of the reductase activity by EDTA. Higher concentrations of iodide inhibit the reductase activity by EDTA. Nitrite, an electron donor, acts similarly to iodide. Both EDTA and nitrite competitively inhibit iodide oxidation, indicating that they compete with iodide for the same binding site for electron flow to the haem iron group. However, unlike iodide, EDTA converts compound I, not into the native enzyme, but into a compound absorbing at 416 nm which reduces I+ and then returns to the native form. The apparent equilibrium dissociation constant, KD, for the formation of the EDTA-HRP complex (15 mM) is doubled in the presence of iodide, indicating interference with EDTA binding by iodide. EDTA binds away from the haem iron centre and not through intramolecular Ca2+. The pH-dependence of EDTA binding indicates that an ionizable group of the enzyme with pKa 5.8, presumably a distal histidine, controls the binding. The data suggest that iodide competes with EDTA for compound I and modulates the iodine reductase activity by limiting the formation of the 416 nm-absorbing active compound.

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