Notomastus lobatus chloroperoxidase and Amphitrite ornata dehaloperoxidase both contain histidine as their proximal heme iron ligand.
Two novel heme-containing peroxidases, one able to incorporate halogens into aromatic substrates and the other able to remove them, have recently been isolated from marine sources and initially characterized by Chen et al. [(1991) J. Biol. Chem. 266, 23909-23915; (1996) J. Biol. Chem. 271, 4609-4612]. The haloperoxidase Notomastus lobatus chloroperoxidase (NCPO) is unusual in requiring a flavoprotein component for peroxidase activity. The dehaloperoxidase (DHP), isolated from Amphitrite ornata, is the only heme-containing peroxide-dependent dehalogenase known to be capable of removing halogens including fluorine. Both enzymes are also quite atypical in that the molecular weights of their heme-containing subunits are less than 16,000, approximately one-half to one-fifth the size of typical heme-containing peroxidases. Interestingly, we have also found that both enzymes are isolated in their oxyferrous states even though all protein purification was done in the absence of any reductant. In the present study, we have examined these two enzymes with magnetic circular dichroism and UV-visible absorption spectroscopy in order to determine the identity of their proximal heme iron ligand. Four derivatives of each enzyme, cyanoferric, deoxyferrous, oxyferrous, and (carbonmonoxy)ferrous, have been examined and spectroscopically compared to parallel derivatives of myoglobin, a well-studied histidine-ligated heme protein. The spectra observed for each derivative of the two new enzymes are very similar to each other and, in turn, to the spectra of the same derivatives of myoglobin. We conclude that both new heme enzymes contain histidine as their proximal heme iron ligand. This makes NCPO the first histidine-ligated heme-containing peroxidase capable of chlorinating halogen acceptor substrates using chloride as the halogen donor. Further, the novel reactivity of DHP is not the result of an unusual proximal ligand. The present results with NCPO and DHP challenge the current dogma of how heme-containing peroxidases function: one chlorinates substrates without having a thiolate proximal ligand, and the other both oxygenates and dehalogenates haloaromatics and yet has a histidine proximal ligand like numerous other peroxidases that are not capable of such a combined reactivity.
Roach
M P
MP
Department of Chemistry and Biochemistry, School of Medicine, University of South Carolina, Columbia 29208, USA.
Chen
Y P
YP
Woodin
S A
SA
Lincoln
D E
DE
Lovell
C R
CR
Dawson
J H
JH
D000818:Animals; D001665:Binding Sites; D002711:Chloride Peroxidase / Q000378:metabolism*; D006418:Heme / Q000378:metabolism*; D006454:Hemoglobins; D006639:Histidine / Q000378:metabolism*; D007501:Iron / Q000378:metabolism*; D008024:Ligands; D010544:Peroxidases / Q000378:metabolism*; D011077:Polychaeta / Q000201:enzymology*
D006454:Hemoglobins; D008024:Ligands; D006418:Heme; D006639:Histidine; D007501:Iron; C098661:DHP I dehaloperoxidase; D010544:Peroxidases; D002711:Chloride Peroxidase