Elongation factor-2 kinase: immunological evidence for the existence of tissue-specific isoforms.
eEF-2 kinase is a ubiquitous Ca2+/calmodulin-dependent protein kinase that is specific for protein synthesis elongation factor-2 (eEF-2). This study describes an improved procedure for the purification of eEF-2 kinase from rabbit reticulocyte lysate. The eEF-2 kinase preparation was used to raise polyclonal antibodies, which immunoprecipitated eEF-2 kinase protein and activity from rabbit reticulocyte lysate. The antibodies recognized a single 103 kDa band in extracts from several cell lines including NIH 3T3, PC12, C6 glioma, HeLa, and MCF-7 breast carcinoma. However, there was no immunoreactivity in extracts of rabbit or bovine liver or rabbit kidney despite the presence of abundant eEF-2 kinase activity in these tissues. Exposure of PC12 cells to nerve growth factor (NGF) resulted in rapid down-regulation of eEF-2 kinase activity and a decrease in immunoreactivity. After 24 h of incubation with NGF, the activity of the kinase recovered to 80% of initial values. In contrast, the immunoreactivity of eEF-2 kinase continued to decrease. These data suggest that tissue-specific isoforms of eEF-2 kinase may exist and that these isoforms may be regulated by growth factors.
Hait
W N
WN
Department of Pharmacology, University of Medicine and Dentistry of New Jersey/Robert Wood Johnson Medical School, Piscataway 08854, USA.
Ward
M D
MD
Trakht
I N
IN
Ryazanov
A G
AG
D016475:3T3 Cells; D000818:Animals; D000906:Antibodies / Q000276:immunology*; D000918:Antibody Specificity; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000032:analysis* / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism; D002417:Cattle; D002847:Chromatography, Agarose; D004591:Electrophoresis, Polyacrylamide Gel; D054736:Elongation Factor 2 Kinase; D006801:Humans; D007124:Immunoenzyme Techniques; D007527:Isoenzymes / Q000032:analysis* / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism; D051379:Mice; D009414:Nerve Growth Factors / Q000494:pharmacology; D009928:Organ Specificity; D016716:PC12 Cells; D011817:Rabbits; D051381:Rats; D014407:Tumor Cells, Cultured
D000906:Antibodies; D007527:Isoenzymes; D009414:Nerve Growth Factors; C517689:EEF2K protein, human; C517691:Eef2k protein, rat; D017871:Calcium-Calmodulin-Dependent Protein Kinases; C517690:Eef2k protein, mouse; D054736:Elongation Factor 2 Kinase