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Histidine modifying agents abolish pyruvate dehydrogenase kinase activity
B P Mooney1, N R David, J J Thelen
1Biochemistry Department, University of Missouri, Columbia, Missouri 65211, USA.
Biochemical and Biophysical Research Communications
|January 13, 2000
Summary
Pyruvate dehydrogenase kinase (PDK) activity, crucial for regulating cellular energy, requires specific histidine residues. Inhibiting these residues with chemical reagents blocks PDK
Area of Science:
- Biochemistry
- Molecular Biology
- Plant Science
Background:
- Pyruvate dehydrogenase kinase (PDK) regulates the pyruvate dehydrogenase complex (PDC) by phosphorylating its E1alpha subunit.
- Sequence analysis suggests PDKs share mechanistic similarities with prokaryotic 2-component histidine kinases.
- Histidine kinases typically involve autophosphorylation of a histidine residue followed by phosphotransfer to an aspartate residue.
Purpose of the Study:
- To investigate the role of histidine residues in the catalytic activity of Arabidopsis thaliana PDK.
- To validate the proposed mechanistic link between PDKs and prokaryotic histidine kinases.
Main Methods:
- Recombinant Arabidopsis thaliana PDK was treated with histidine-directed chemical reagents: diethyl pyrocarbonate (DEPC) and dichloro-(2,2 :6 , 2"-terpyridine)-platinum(II) dihydrate.
- The effects of these treatments on PDK autophosphorylation and its ability to inactivate PDC were assessed.
Main Results:
- Treatment with DEPC and the platinum compound significantly inhibited PDK autophosphorylation.
- DEPC treatment abolished PDK's capacity to trans-phosphorylate and consequently inactivate the PDC.
Conclusions:
- These findings experimentally validate the prediction that histidine residues are essential for PDK activity.
- The results support the mechanistic relationship between plant PDKs and prokaryotic 2-component histidine kinases.