Biochemistry of coproporphyrin ferrochelatases

Heme is essential for the survival of most bacteria. Gram-positive organisms produce heme in a way that is fundamentally different from the biosynthetic pathway used by Gram-negative organisms or even mammals. Many mechanistic questions relating to this heme biosynthetic pathway, which was only described a few years ago, are currently still open. In this project, the enzyme called "coproporphyrin ferrochelatase" is being studied in detail to elucidate structure-function relationships. Ferrochelatases incorporate ferrous iron atoms into a porphyrin ring and have been investigated in protein biochemical research for several decades. However, the studies have always been carried out with a substrate that, as has just been shown, is not physiologically relevant, namely protoporphyrin IX. The correct substrate, however, is coproporphyrin III, which has two more propionate groups than protoporphyrin IX and therefore has different binding properties. Coproporphyrin ferrochelatase from the Gram-positive pathogenic bacterium Listeria monocytogenes and variants, in which individual amino acid residues are exchanged at catalytically important sites, are produced recombinantly in Escherichia coli and purified to obtain a highly pure protein for biochemical and biophysical characterizations. Using several sophisticated spectroscopic, structural and biochemical analysis methods, the natural wild-type form of coproporphyrin ferrochelatase is studied in detail and compared with the variants that are punctually altered. From these studies essential conclusions can be drawn about the mode of action of this enzyme and can be linked to its structural properties. Knowledge of the reaction mechanism of copropophryin ferrochelatases is necessary to design further studies that will attempt to inhibit enzymatic activity. A substance that can specifically inhibit the heme biosynthesis pathway of pathogenic Gram-positive bacteria is a promising starting point for the development of urgently needed novel antibiotics.