title: Histidine modifying agents abolish pyruvate dehydrogenase kinase activity.
abstract: Pyruvate dehydrogenase kinase (PDK) specifically phosphorylates the E1alpha subunit of the pyruvate dehydrogenase complex (PDC). Sequence analysis of cloned PDKs led to the proposal that they are mechanistically related to prokaryotic 2-component His-kinases. The reaction mechanism of protein His-kinases involves autophosphorylation of a specific His residue followed by phosphotransfer to an Asp residue. Treatment of recombinant Arabidopsis thaliana PDK with the His-directed reagents diethyl pyrocarbonate (DEPC) and dichloro-(2,2':6', 2"-terpyridine)-platinum(II) dihydrate led to a marked inhibition of autophosphorylation. In addition, DEPC treatment abolished the ability of PDK to trans-phosphorylate and inactivate PDC. These results validate the prediction that PDKs require His residues for activity.
authors.affiliation: Biochemistry Department, University of Missouri, Columbia, Missouri 65211, USA.
authors.forename: B P
authors.initials: BP
authors.lastname: Mooney
authors.forename: N R
authors.initials: NR
authors.lastname: David
authors.forename: J J
authors.initials: JJ
authors.lastname: Thelen
authors.forename: J A
authors.initials: JA
authors.lastname: Miernyk
authors.forename: D D
authors.initials: DD
authors.lastname: Randall
mesh_terms: D017360:Arabidopsis / Q000201:enzymology / Q000235:genetics; D004047:Diethyl Pyrocarbonate / Q000494:pharmacology; D006639:Histidine / Q000378:metabolism*; D000071677:Histidine Kinase; D009944:Organoplatinum Compounds / Q000494:pharmacology; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases; D030481:Pyruvate Dehydrogenase (Lipoamide); D000081382:Pyruvate Dehydrogenase Acetyl-Transferring Kinase; D011768:Pyruvate Dehydrogenase Complex / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism
chemical_list: D009944:Organoplatinum Compounds; D047428:Protein Kinase Inhibitors; D000081382:Pyruvate Dehydrogenase Acetyl-Transferring Kinase; D011768:Pyruvate Dehydrogenase Complex; D011993:Recombinant Fusion Proteins; D006639:Histidine; D030481:Pyruvate Dehydrogenase (Lipoamide); C065640:pyruvate dehydrogenase E1alpha subunit; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D000071677:Histidine Kinase; D004047:Diethyl Pyrocarbonate