Novel tetrapyrrole biosynthetic routes in prokaryotes: Structure and function of enzymes for the biosynthesis of heme d1 in denitrifying bacteria and heme in archaea

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Cyclic tetrapyrroles such as heme and chlorophyll fulfil important biological functions and are involved in processes like aerobic and anaerobic respiration or photosynthesis. Heme d1 is another cyclic tetrapyrrole, which is important for bacterial denitrification and thus for the global nitrogen cycle. The biosynthesis pathways of heme in sulfate-reducing bacteria and archaea and of heme d1 in denitrifying bacteria are related and were elucidated only recently. One reaction occurring in both pathways is the decarboxylation of two acetate groups of siroheme to the corresponding methyl groups of didecarboxysiroheme catalyzed by the enzyme siroheme decarboxylase. The Radical SAM enzyme AhbD catalyzes the last reaction of the siroheme-dependent biosynthesis route to heme, namely the oxidative decarboxylation of two propionate groups of the intermediate iron-coproporphyrin III yielding the two vinyl groups of heme. Within this project, the catalytic mechanisms of the siroheme decarboxylase and of the heme synthase AhbD will be investigated. In the case of siroheme decarboxylase, there are different isoforms of the enzyme that catalyze either only one or both of the two decarboxylation reactions. Therefore, one aim of this project is to identify the respective amino acid residues, which are involved in substrate binding and catalysis, and to identify the determinants responsible for the substrate and reaction specificity of the different siroheme decarboxylase isoforms. Concerning AhbD, the product of the decarboxylation event will be identified and the radical mechanism of the enzyme will be studied by EPR spectroscopy. Another aim is the determination of the three-dimensional structure of AhbD in particular in complex with its substrate.