14-3-3-ALPHA AND 14-3-3-DELTA ARE THE PHOSPHORYLATED FORMS OF RAF-ACTIVATING 14-3-3-BETA AND 14-3-3-ZETA - IN-VIVO STOICHIOMETRIC PHOSPHORYLATION IN BRAIN AT A SER-PRO-GLU-LYS MOTIF

The 14-3-3 protein family has received considerable attention recently in the literature, because of the finding that beta and zeta isoforms interact with and activate Raf. We had previously shown that these 14-3-3 isoforms also exist as phosphorylated forms in mammalian and avian brain. The presence of this modification enhances the activity of 14-3-3 as an inhibitor of protein kinase C nearly 2-fold. In this report we show by a combination of electrospray mass spectrometry and protein microsequencing that alpha and delta are in vivo post-translationally modified forms of beta and zeta, respectively, and the site of phosphorylation, serine 185, is in a consensus sequence motif for proline-directed kinases.