Cloning and biochemical characterization of a plant protein kinase that phosphorylates serine, threonine, and tyrosine.
Phosphorylation of proteins on serine, threonine, or tyrosine residues represents an important biochemical mechanism to regulate the activity of enzymes and is used in many cellular processes. In animals, protein serine/threonine and protein tyrosine kinases are known to perform essential roles in many pathways that transmit external stimuli from the cell surface to the cell inferior and the nucleus. In plants, although an increasing number of protein serine/threonine kinases have been cloned, the existence of protein tyrosine kinases remains yet to be demonstrated. Here, we report the cloning and biochemical characterization of a plant protein kinase, Arabidopsis dual specificity kinase 1 (ADK1), using a functional screening method, namely by screening an Arabidopsis expression library with antiphosphotyrosine antibodies. Four independent cDNA clones that define a polypeptide of 319 amino acids length with homology to protein kinases were identified in this screen. Phosphoamino acid analysis of the autophosphorylated kinase shows that ADK1 phosphorylates serine, threonine, and tyrosine. Using poly (Glu/Tyr) as a substrate, we confirm that ADK1 is capable of phosphorylating tyrosine residues.
Ali
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Laboratory of Plant Molecular Biology, Rockefeller University, New York, New York 10021.
Halfter
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U
Chua
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NH
D017360:Arabidopsis / Q000201:enzymology; D029681:Arabidopsis Proteins; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers / Q000737:chemistry; D008969:Molecular Sequence Data; D010768:Phosphoserine / Q000378:metabolism; D010769:Phosphothreonine / Q000378:metabolism; D019000:Phosphotyrosine; D010940:Plant Proteins / Q000378:metabolism; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D014443:Tyrosine / Q000031:analogs & derivatives / Q000378:metabolism
D029681:Arabidopsis Proteins; D017931:DNA Primers; D010940:Plant Proteins; D011994:Recombinant Proteins; D010769:Phosphothreonine; D010768:Phosphoserine; D019000:Phosphotyrosine; D014443:Tyrosine; D011494:Protein Kinases; C090910:ADK1 protein, Arabidopsis; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases