Submitted to: European Journal of Biochemistry
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/15/2002
Publication Date: 5/1/2002
Citation: TOVAR-MENDEZ, A., MIERNYK, J.A., RANDALL, D.D. HISTIDINE MUTAGENESIS OF ARABIDOPSIS THALIANA PYRUVATE DEHYDROGENASE KINASE. EUROPEAN JOURNAL OF BIOCHEMISTRY. 2002. v. 269. p. 2601-2606. Interpretive Summary: Respiration is the use of energy by living cells to do work. Both growth and reproduction are directly coupled to rates of respiration. As a result, respiration must be carefully controlled or wasted energy would decrease crop yields and reduce agricultural productivity. The control of respiration in plant cells is a subject of ongoing study. A protein that is important in the regulation of respiration was isolated from the model plant, Arabidopsis thaliana, and studied. Comparisons were made with closely related proteins from animals and microbes in order to predict characteristics that might be important in control of respiration. Based upon these comparisons, a mechanism of action for the protein was predicted. A method was developed to test this prediction. Results indicated that the predicted mechanism was incorrect, and an alternative explanation was developed. This information will be important to researchers in their attempts to increase agricultural productivity by altering the control of plant cell respiration, and to other plant scientists who will try to design more efficient crop plants through either classical breeding or biotechnology.
Technical Abstract: Pyruvate dehydrogenase kinase (PDK) is the primary regulator of flux through the mitochondrial pyruvate dehydrogenase complex (PDC). Analysis of the primary amino acid sequences of PDK from various sources reveals that these enzymes include the five domains characteristic of prokaryotic two-component His-kinases. This despite the fact that PDK exclusively phosphorylates Ser residues in the E1alpha subunit of the PDC. This seeming contradiction might be resolved if the PDK-catalyzed reaction employed a phospho-His intermediate. The results from pH stability studies of autophosphorylated Arabidopsis thaliana PDK did not provide any support for a phospho-His intermediate. Furthermore, site-directed mutagenesis of the two most likely phosphotransfer His residues (H121 and H168) did not abolish either PDK autophosphorylation or the ability to transphosphorylate E1alpha. Thus, PDK is a unique type of protein kinase having a His-kinase-like sequence but Ser-kinase activity.