Cloning and expression of cDNA encoding protein synthesis elongation factor-2 kinase.
A cDNA from rat skeletal muscle encoding calcium/calmodulin-dependent eukaryotic elongation factor-2 kinase (eEF-2K) has been cloned and sequenced, and the amino acid sequence of the protein has been deduced. The kinase is composed of 724 amino acids and has a predicted molecular mass of 81,499 Da. The cDNA was judged to be full-length, as the protein, expressed in rabbit reticulocyte lysate or wheat germ extract, migrated upon SDS-PAGE with the same apparent molecular weight as the purified kinase and possessed eEF-2K activity. eEF-2K contains all of the 12 catalytic subdomains present in the majority of protein kinases, but they are atypical and display only limited homology with other kinases. A putative calmodulin-binding domain is present C-terminal to the catalytic domain as is a putative pseudosubstrate sequence. Two antipeptide antibodies raised against sequences derived from a partial rabbit cDNA clone, cross-reacted with purified eEF-2K, and one also immunoprecipitated eEF-2K activity from cell extracts. Northern blot analysis demonstrated that eEF-2K mRNA is expressed in a number of different tissues and that it may exist in multiple forms.
Redpath
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Department of Biochemistry, University of Leicester, University Road, Leicester LE1 7RH, United Kingdom.
Price
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Proud
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D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D017173:Caenorhabditis elegans / Q000201:enzymology; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification; D003001:Cloning, Molecular; D016384:Consensus Sequence; D017124:Conserved Sequence; D017931:DNA Primers; D018076:DNA, Complementary; D054736:Elongation Factor 2 Kinase; D015870:Gene Expression; D007527:Isoenzymes / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D008969:Molecular Sequence Data; D008970:Molecular Weight; D018482:Muscle, Skeletal / Q000201:enzymology*; D020652:Peptide Elongation Factor 2; D010445:Peptide Elongation Factors / Q000737:chemistry / Q000378:metabolism; D016133:Polymerase Chain Reaction; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000096:biosynthesis; D011817:Rabbits; D051381:Rats; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D012156:Reticulocytes / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014158:Transcription, Genetic
D017931:DNA Primers; D018076:DNA, Complementary; D007527:Isoenzymes; D020652:Peptide Elongation Factor 2; D010445:Peptide Elongation Factors; D012333:RNA, Messenger; D011994:Recombinant Proteins; C517691:Eef2k protein, rat; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D054736:Elongation Factor 2 Kinase