Ferrochelatase yeast

Hie expression of recombinant mammalian liver ferrochelatases in Escherichia coli [4, 5], allowed the production of increased amounts of the enzyme and led to the unexpected discovery of a [2Fe-2S] cluster in the mammalian enzymes (mouse [6] and human [7]). The large amoimts of protein made available using heterologous over-producing systems allowed detailed spectroscopic studies on the cluster-containing ferrochelatases. Yeast Saccharomyces cerevisiae) ferrochelatase, which is devoid of an iron-sulfur cluster, has also been extensively studied and several site-directed mutants have been produced [8], making it a powerful model system to study the mechanism of this enzyme. 

Ferrochelatase (protoheme ferro-lyase)401 403 inserts Fe2+ into protoporphyrin IX to form heme. The enzyme is found firmly bound to the inner membrane of mitochondria of animal cells, chloroplasts of plants, and chromatophores of bacteria. While Fe2+ is apparently the only metallic ion ordinarily inserted into a porphyrin, the Zn2+ protoporphyrin chelate accumulates in substantial amounts in yeast, and Cu2+-heme complexes are known (p. 843). Ferrochelatase, whose activity is stimulated by Ca2+, appears to be inhibited by lead ions, a fact that may account for some of the acute toxicity of lead.404... 

The combined use of spectroscopy and protein engineering techniques provides a unique approach for the study of the mechanism of ferrochelatase and in the elucidation of the role of its intriguing [2Fe-2S] cluster. Mossbauer studies of kinetic intermediates and fluorimetric and resonance Raman studies of both mammalian and yeast (i.e. devoid of the [2Fe-2S] cluster) ferrochelatases, reacted with porphyrin or porphyrin analogs, should be undertaken. Determination of the three-dimensional structures of the complexes of ferrochelatase with substrate or inhibitors will also be of crucial importance. 

A [2Fe-2S] cluster is also present in mammalian ferrochelatase, the terminal enzyme of the heme biosynthetic pathway, and it appears to be the determinant for catalysis as reviewed by Franco et al. (Chapter 3). Nevertheless, the specific role of this cluster in the enzyme s stability and function remains to be established. The search for the function of this cluster becomes even more challenging as bacterial, yeast and plant ferrochelatases do not possess it, although they perform the same catalytic task. 

Frataxin involvement in heme synthesis was suggested due to the striking phenotype of frataxin deficient yeast mutants almost completely lacking heme biosynthetic activity while retaining zinc chelatase activity 4). A direct interaction between frataxin and ferrochelatase was originally shown with the... 

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