Synthesis and reactivity studies of a manganese 'microperoxidase' containing b-type heme

Mn(III)protoporphyrin IX-6(7)-gly-gly-his methyl ester (MnGGH) has been prepared by condensation of glycyl-glycyl-L-histidine methyl ester with the propionic side chains of Mn(III)protoporphyrin IX. It was characterised by mass spectrometry and UV/VIS spectroscopy. Stopped-flow spectrophotometry was used to study the reaction of the Mn 'microperoxidase' with hydrogen peroxide. The formation of active intermediates analogous to previously described metal-hydroperoxo (compound 0) and metal-oxo (compound I) intermediates of the 'natural' Fe(III) microperoxidase-8 and Mn(III) microperoxidase-8 was observed. The rate of formation of the MnGGH-based compound I analogue was found to increase dramatically with increasing pH. A steady-state kinetic analysis of the catalytic peroxidase activity of MnGGH towards K4[Fe(CN)6], L-tyrosine methyl ester, o-dianisidine, o-methoxyphenol and ascorbic acid showed that the peroxidase reaction proceeds via the formation of a microperoxidase-substrate complex followed by electron transfer from the substrate to the metal. The reactivity of MnGGH depends on the size and hydrophobicity of the substrate, and these properties appear to influence the rate of the electron transfer, which is the rate-limiting step for the whole process. MnGGH showed higher reactivity towards reducing substrates than its Fe(iii) analogue.