Protein synthesis in rabbit reticulocyte lysates is regulated by heme. In heme deficiency, a heme regulated protein kinase (HRI) is activated that phosphorylates initiation factor eIF-2. Consequently, eIF-2 is inactivated. Results described in this report show that HRI exists in crude and highly purified preparations in two forms; a high molecular weight component which sediments at a sedimentation co-efficient of 14-15S and a previously described 5.8S component (Ranu, R. S. and London, I. M. (1976) Proc. Natl. Acad. Sci. USA 73, 4349-4353). The 14-15S HRI selfphosphorylates poorly and undergoes dissociation into the 5.8S component via an intermediate of 8.5-9S. The 5.8S HRI, on weight basis, is about 5-10 times more active than the 14-15S HRI. In addition, a phosphoprotein phosphatase has been detected in lysates that dephosphorylates selfphosphorylated HRI. This observation suggests that phosphate on HRI turns over. These findings may be relevant to the mechanism of activation and inactivation of HRI in the absence and presence of heme in situ.