ISOLATION OF A CDNA-ENCODING CHLOROPLAST FERROCHELATASE FROM ARABIDOPSIS-THALIANA BY FUNCTIONAL COMPLEMENTATION OF A YEAST MUTANT

Ferrochelatase catalyzes the insertion of ferrous iron into protoporph yrin IX to form protoheme. It is located in the mitochondria in all eu karyotes and is also found in plastids in plants. Although it has been purified from animals and microorganisms, and genes for it isolated a nd characterized, very little is known about plant ferrochelatases. We have isolated a cDNA for ferrochelatase from the higher plant Arabido psis thaliana by functional complementation of a mutant of Saccharomyc es cerevisiae defective in this enzyme. The cDNA encodes a protein of 52 kDa, which has 25-35% sequence similarity to ferrochelatases from o ther organisms. There is an N-terminal extension of about 65 residues, which is almost certainly the chloroplast transit peptide, since the precursor protein, transcribed and translated in vitro, is efficiently imported and processed to the mature size by isolated pea chloroplast s. In contrast, the precursor was not processed by mitochondrial proce ssing peptidase activity, nor could import into isolated yeast mitocho ndria be demonstrated conclusively, although, presumably, in the rescu ed yeast mutant, at least some of the Arabidopsis ferrochelatase must be present in the mitochondria. A single transcript the same size as t he cDNA was detected in both Arabidopsis leaves and roots, although th e amount of message was greater in the photosynthetic tissue. Southern analysis suggests that there is a single gene for chloroplast ferroch elatase in Arabidopsis.