Ferrochelatase is the terminal enzyme of the heme biosynthetic pathway in all cells. It catalyzes the insertion of ferrous iron into protoporphyrin IX, yielding heme. In eukaryotic cells, ferrochelatase is a mitochondrial inner membrane-associated protein with the active site facing the matrix. Decreased values of ferrochelatase activity in all tissues are a characteristic of patients with protoporphyria. Point-mutations in the ferrochelatase gene have been recently found to be associated with certain cases of erythropoietic protoporphyria. During the past four years, there have been considerable advances in different aspects related to structure and function of ferrochelatase. Genomic and cDNA clones for bacteria, yeast, barley, mouse, and human ferrochelatase have been isolated and sequenced. Functional expression of yeast ferrochelatase in yeast strains deficient in this enzyme, and expression in Escherichia coli and in baculovirus-infected insect cells of different ferrochelatase cDNAs have been accomplished. A recently identified (2Fe-2S) cluster appears to be a structural feature shared among mammalian ferrochelatases. Finally, functional studies of ferrochelatase site-directed mutants, in which key amino acids were replaced with residues identified in some cases of protoporphyria, will be summarized in the context of protein structure.