Three phosphorylation sites in elongation factor 2

[摘要]

Elongation factor 2 (EF-2) of rabbit reticulocytes was phosphorylated in vitro by incubation with partially purified EF-2 kinase and (γ³²P)ATP. After exhaustive tryptic hydrolysis 4 phosphopeptides were revealed by two-dimensional peptide mapping. The phosphopeptides were isolated by high performance liquid chromatography and sequenced. A comparison of the primary structure of the phosphopeptides with that of EF-2 showed that all 4 phosphopeptides originated from one region of EF-2 located near the N-terminus that contains 3 threonine residues: Thr-53, Thr-56, Thr-58. A direct estimation of localization of radioactive phosphate in the phosphopeptides demonstrated that all the enumerated threonine residues in EF-2 can be phosphorylated in vitro.

[学科分类] 生物化学/生物物理

[关键词] Elongation factor 2;Phosphorylation;EF-2 kinase;Tryptic phosphopeptide;EF-1;EF-2;elongation factor 1;elongation factor 2;DEAE-cellulose;diethylaminoethyl cellulose;TPCK;L-1-tosyl-amino-2-phenylethyl chloromethyl ketone;SDS;sodium dodecyl sulfate;Hepes;4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid;DTT;dithiothreitol;TCA;trichloroacetic acid;RP-HPLC;reversed-phase high performance liquid chromatography;TFA;trifluoroacetic acid;EDTA;ethylenediaminetetraacetic acid;PTH;2-phenyl-5-thio-hydantoin;MES;2-(N-morpholino) ethanesulfonic acid;ATZ;2-anilino-5-thiazolinone