An Investigation of Peroxidase Activity in Biomimetic and Biological Systems

Sammanfattning: In order to create a heme environment which permits biomimicry of heme-containing plant peroxidases and investigate the role played by the vinyl side chains of the porphyrin ring on the stability and, possibly, catalytic activity of model compounds, a number of new synthetic microperoxidases -– hemin-6(7)-gly-his methyl ester (HGH) and hemin-6(7)-gly-gly-his methyl ester (HGGH) and hemin-6,7-bis(gly-gly-his methyl ester) (H2GGH) as well as mesohemin-6(7)-gly-gly-his methyl ester (MGGH) and deuterohemin-6(7)-gly-gly-his methyl ester (DGGH) have been prepared by condensation of a peptide residue with the propionic side chains of hemin, mesohemin and deuterohemin, respectively. Reactivity studies towards different organic and inorganic substrates showed that not only five- but also six-coordinate hemin peptide complexes catalyze the oxidation of substrates by H2O2. The HGGH complex showed higher catalytic activity than HGH due to a less strained structure provided by the longer peptide arm. Microperoxidases based on meso- and deuterohemin (MGGH and DGGH, respectively) exhibited at least one order of magnitude higher reactivity relative to their hemin-based analogues. However, computational modelling showed that the relative energy differences between the different hemins are small, suggesting that more subtle factors (e.g. properties of reaction media) rather that only electronic properties of substituents of the porphyrin ring determine the reactivity order of the hemin peptide complexes studied. In general, the hemin-peptide complexes showed relatively low peroxidase reactivity. On the contrary, manganese(III) microperoxidase (MnGGH) showed relatively high reactivity in peroxidase-type reactions. Rate constants for Compound I formation obtained by fast kinetics studies as well as steady-state rate constants for the reaction of MnGGH with the substrates were comparable to these observed for manganese microperoxidase-8 (MnMP-8). The higher peroxidase reactivity of MnGGH as compared to its iron(III) analogue, HGGH, may be attributed to the higher operational stability of the manganese complexes as well as higher stability of Mn(III) oxo intermediates. An investigation of the peroxidase reactivity of bacterial hemoglobin from Vitreoscilla stercoraria (VHb) showed for the first time that VHb does exhibit peroxidase catalytic activity which for certain substrates is comparable with the activity of horseradish peroxidase. VHb also showed unusual substrate specificity (for peroxidases); i.e. good activity was observed only for substrates with more than one hydrogen bond donor/acceptor. In order to create a heme environment which permits biomimicry of heme-containing plant peroxidases and investigate the role played by the vinyl side chains of the porphyrin ring on the stability and catalytic activity of model compounds, a number of new synthetic microperoxidases – hemin-6(7)-gly-his methyl ester (HGH), hemin-6(7)-gly-gly-his methyl ester (HGGH) and hemin-6,7-bis(gly-gly-his methyl ester) (H2GGH) as well as mesohemin-6(7)-gly-gly-his methyl ester (MGGH) and deuterohemin-6(7)-gly-gly-his methyl ester (DGGH) have been prepared by condensation of a peptide residue with the propionic acid side chains of hemin, mesohemin and deuterohemin, respectively. Reactivity studies towards different organic and inorganic substrates showed that not only five- but also six-coordinate hemin peptide complexes catalyze the oxidation of substrates by H2O2. The HGGH complex showed higher catalytic activity than HGH, possibly due to a less strained structure provided by the longer peptide arm. Microperoxidases based on meso- and deuterohemin (MGGH and DGGH, respectively) exhibited at least one order of magnitude higher reactivity relative to their hemin-based analogues. However, computational modelling showed that the relative energy differences between the different hemins are small, suggesting that more subtle factors (e.g. properties of reaction media) rather that only electronic properties of the substituents of the porphyrin ring determine the reactivity order of the hemin peptide complexes studied. Manganese(III) microperoxidase (MnGGH) showed relatively high reactivity in peroxidase-type reactions. Rate constants for Compound I formation obtained by fast kinetics studies as well as steady-state rate constants for the reaction of MnGGH with substrates were comparable to these observed for manganese microperoxidase-8. The higher peroxidase reactivity of MnGGH as compared to its iron(III) analogue, HGGH, may be attributed to the higher operational stability of the manganese complex as well as higher stability of Mn(III) oxo intermediates. An investigation of the peroxidase reactivity of the bacterial hemoglobin from Vitreoscilla stercoraria (VHb) showed for the first time that VHb does exhibit peroxidase catalytic activity which for certain substrates is comparable with the activity of horseradish peroxidase. VHb also showed unusual substrate specificity (for peroxidases); i.e. good activity was observed only for substrates with more than one hydrogen bond donor/acceptor.