Spectroscopic studies on lacquer peroxidase from Rhus vernicifera and its derivatives

Abstract

In order to probe the active site of lacquer peroxidase (LAP) from Rhus vernicifera, electronic absorption and magnetic circular dichroism (MCD) studies of the enzyme with and without various exogenous ligands (NO, CO, and CN −) have been carried out at room temperature. The absorption and MCD spectra of native LAP are quite similar to those of horseradish peroxidase (HRP), indicating that the enzyme has a heme with the fifth and the sixth coordination sites occupied by histidine imidazole and an exogenously exchangeable ligand (H 2O), respectively. Moreover, the superhyperfine structure of the electron paramagnetic resonance (EPR) signal of NO-ferrous LAP provides strong support for the fifth imidazole ligand. A comparison of the Soret MCD band of native LAP with that of Hg(II)-treated LAP demonstrated that an equilibrium mixture of high-spin heme (major species) and low-spin heme (minor species) is present in the enzyme at room temperature. All of the derivatives of NO, CO, and CN − exhibit electronic and MCD spectra of low-spin complexes; similar spectral features are found for the corresponding derivatives of HRP and myoglobin (Mb). The treatment of LAP with H 2O 2 gives rise to stable compound II at room temperature, while compound I is too unstable to be spectroscopically observed.