The synthesis of murine ferrochelatase in vitro and in vivo

Abstract

Ferrochelatase (protohaem ferro-lyase, EC 4.99.1.1), the terminal enzyme of the haem-biosynthetic pathway, is an integral membrane protein of the mitochondrial inner membrane. When murine erythroleukaemia cells are labelled in vivo with [35S]methionine, lysed, and the extract is immunoprecipitated with rabbit anti-(mouse ferrochelatase) antibody, a protein of M(r) 40,000 is isolated. However, when isolated mouse RNA is translated in a cell-free reticulocyte extract, a protein of M(r) 43,000 is isolated. Incubation of this M(r) 43,000 protein with isolated mitochondria resulted in processing of the M(r) 43,000 precursor to the M(r) 40,000 mature-sized protein. Addition of carbonyl cyanide m-chlorophenylhydrazone and/or phenanthroline inhibits this processing. These data indicate that ferrochelatase, like most mitochondrial proteins, is synthesized in the cytoplasm as a larger precursor and is then translocated and processed to a mature-sized protein in an energy-required step.