Chemical and kinetic evidence for an essential histidine residue in the electron transfer from aromatic donor to horseradish peroxidase compound I.
Horseradish peroxidase, when incubated with diethyl pyrocarbonate (DEPC), a histidine-specific reagent, shows time-dependent inactivation to oxidize aromatic electron donor, guaiacol. The inactivation follows pseudo-first order kinetics with a second order rate constant of 0.67 min-1 M-1. The pH dependence of inactivation shows an inflection point at 6.02, indicating histidine derivatization by DEPC. A difference spectrum of modified versus native enzyme shows a peak at 244 nm for N-carbethoxyhistidine that is diminished by hydroxylamine. Stoichiometric studies indicate that out of 2 histidine residues modified, one is responsible for inactivation. A plot of log reciprocal half-time of inactivation against log DEPC concentration suggests that only 1 histidine is essential. From the computer-stimulated structure of horseradish peroxidase, we tentatively suggest that this critical histidine is most likely distal histidine 42. Binding studies show that this histidine is not involved in guaiacol binding. Modified enzyme forms compound I with H2O2 but not compound II, suggesting a block of electron transfer process. Modified compound I cannot oxidize guaiacol as evidenced by the absence of donor-induced spectral shift from 408 nm, suggesting a block of electron transfer from bound donor to compound I. We suggest that this tentatively identified distal histidine controls aromatic donor oxidation by regulating electron transport without affecting donor binding or compound I formation.[1]References
- Chemical and kinetic evidence for an essential histidine residue in the electron transfer from aromatic donor to horseradish peroxidase compound I. Bhattacharyya, D.K., Bandyopadhyay, U., Banerjee, R.K. J. Biol. Chem. (1993) [Pubmed]
Annotations and hyperlinks in this abstract are from individual authors of WikiGenes or automatically generated by the WikiGenes Data Mining Engine. The abstract is from MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.About WikiGenesOpen Access LicencePrivacy PolicyTerms of Useapsburg
