Identification of novel phosphorylation sites in the β-subunit of translation initiation factor eIF-2

Initiation factor eIF-2 (a trimer of subunits α, β and γ) attaches the initiator Met-tRNA to the ribosome during the initiation of translation in eukaryotic cells. Both the α and β subunits can be phosphorylated although the sites in the β-subunit have not previously been fully identified. Here we identify the sites at which eIF-2β is phosphorylated in vitro by three well-characterised protein kinases casein kinase-2 (which phosphorylates serine residues-2 and -67), protein kinase C (serine-13) and cAMP-dependent protein kinase (serine-218). This constitutes an essential prerequisite for studying the phosphorylation of eIF-2β in vivo. Indeed, we present evidence that at least one of these sites (serine-67) is phosphorylated in reticulocytes. The major kinase activity against eIF-2β in reticulocyte lysates appears in CK-2 and protein phosphatase-2A is the principal enzyme responsible for dephosphorylation of eIF-2β phosphorylated by this kinase.