Open AccessKinetic Studies of the Reaction of Ferric Soybean Leghemoglobins with Hydrogen Peroxide, Cyanide and Nicotinic Acid
Author(s) -
JOB Dominique,
ZEBA Boukaré,
PUPPO Alain,
RIGAUD Jean
Publication year - 1980
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1980.tb06055.x
Subject(s) - chemistry , acid dissociation constant , reaction rate constant , protonation , dissociation constant , hydrogen peroxide , dissociation (chemistry) , inorganic chemistry , titration , leghemoglobin , photochemistry , heme , medicinal chemistry , organic chemistry , nitrogen , aqueous solution , kinetics , enzyme , ion , root nodule , biochemistry , physics , receptor , quantum mechanics , nitrogen fixation
A kinetic study of the reaction of two soybean leghemoglobins (components a and c) with hydrogen peroxide to form the oxidized compound (leghemoglobin IV) has been carried out over the pH range 2.5–10. Three different ionization processes of leghemoglobins with p K a values of 3, 4.7 ± 0.2 and 8.2 ± 0.1 are required to explain the rate/pH profiles. Protonation of the former group and ionization of the latter cause a decrease in the rate of reaction of the hemoproteins with H 2 O 2 . The results are compared to those obtained for the reactions of plant peroxidases and myoglobin with H 2 O 2 . The results obtained from the kinetic study of cyanide binding to soybean leghemoglobins indicate that CN − is the reactive species. Two ionization processes of leghemoglobins with p K a values of 4.7 ± 0.2 and 8.2 ± 0.1 affect the reaction rates. The association and dissociation rate constants corresponding to nicotinic acid binding to leghemoglobins a and c have been measured over the pH range 2.5–7. The dissociation rate constant is affected by ionization of a group with p K a < 2.5 for both leghemoglobin‐nicotinate complexes. In this pH range the association rate constant is only affected by ionization of a group with p K a value of 4.7 ± 0.2. The analysis of these results shows that both ionization processes corresponding to ring nitrogen atom of the ligand (p K a ∼ 4.9) and to a heme‐linked group (p K a ∼ 4.7 ± 0.2) influence the association rate constant. Furthermore, it appears that in the binding site of leghemoglobins the p K a value corresponding to ionization of the ring nitrogen atom of nicotinic acid is shifted from the normal value of 4.9 to a value < 2.5. This pecularity might explain the exceptional reactivity of leghemoglobins for nicotinic acid, over a large pH range. For both cyanide and nicotinic acid binding reactions, the ionizable group of leghemoglobins with p K a value of 4.7 ± 0.2 seems to act as an electrostatic gate. When this group is deprotonated, it restricts the access of anion ligands to the heme pocket. For all the three reactions studied, leghemoglobin a reacts about twice as fast as leghemoglobin c.