An Essential Role of Glu-243 and His-239 in the Phosphotransfer Reaction Catalyzed by Pyruvate Dehydrogenase Kinase

Tuganova, Alina; Yoder, Marilyn D.; Popov, Kirill M.

doi:10.1074/jbc.m009327200

Cited by 23 publications

(20 citation statements)

References 36 publications

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“…Instead of attacking the ␥-phosphate of ATP with the side chain of the phosphateaccepting histidine in the H-box (41) it uses a glutamate in the N-box as a general base catalyst to activate the phosphateaccepting serine (42). The results presented here indicate that the conserved histidine residue of the H-box is not required for serine autophosphorylation of the ethylene receptors (Fig.…”

Section: Fig 3 Nature Of Phosphorylated Amino Acidsmentioning

confidence: 50%

Autophosphorylation Activity of the Arabidopsis Ethylene Receptor Multigene Family

Moussatche¹,

Klee

2004

Journal of Biological Chemistry

187

157

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Receptors for the gaseous phytohormone ethylene show sequence similarity to bacterial two-component histidine kinases. These receptors are encoded by a multigene family that can be divided into subfamilies 1 and 2. It has been previously shown that a subfamily 1 Arabidopsis thaliana ethylene receptor, ETR1, autophosphorylates in vitro on a conserved histidine residue (1). However, sequence comparisons between the five ethylene receptor family members suggest that subfamily 2 members do not have all the motifs necessary for histidine kinase activity. Further, a tobacco subfamily 2 receptor, NTHK1, autophosphorylates on serines and threonines in vitro (2). Here we show that all five Arabidopsis ethylene receptor proteins autophosphorylate in vitro. We analyzed the nature of the phosphorylated amino acids by acid/base stability and bi-dimensional thin layer electrophoresis and demonstrated that unlike ETR1 all other ethylene receptors autophosphorylate predominantly on serine residues. ERS1, the only other subfamily 1 receptor, is able to phosphorylate on both histidine and serine residues in the presence of Mn 2؉ . However, histidine autophosphorylation is lost when ERS1 is assayed in the presence of both Mg 2؉ and Mn 2؉ , suggesting that this activity may not occur in vivo. Furthermore, mutation of the histidine residue conserved in two-component systems does not abolish serine autophosphorylation, eliminating the possibility of a histidine to serine phosphotransfer. Our biochemical observations complement the recently published genetic data that histidine kinase activity is not necessary for ethylene receptor function in plants and suggest that ethylene signal transduction does not occur through a phosphorelay mechanism.

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Section: Fig 3 Nature Of Phosphorylated Amino Acidsmentioning

confidence: 50%

Autophosphorylation Activity of the Arabidopsis Ethylene Receptor Multigene Family

Moussatche¹,

Klee

2004

Journal of Biological Chemistry

187

157

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“…This model is dissimilar to that used by PDK and BCKDK which phosphorylate serine residues on their target proteins. PDK can to catalyse the transfer of the gphosphate to the side chain of serine on its substrate E1 [18,19]. Glutamate 243 of PDK appears essential in the mechanism, by activating the hydroxyl group of the target serine and thus priming it for interaction with the gphosphate [19,20].…”

Section: Protein Kinases Associated With Metabolism and Respirationmentioning

confidence: 98%

Evidence of undiscovered cell regulatory mechanisms: phosphoproteins and protein kinases in mitochondria

Thomson¹

2002

CMLS, Cell. Mol. Life Sci.

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The finding that mitochondria contain substrates for protein kinases lead to the discovery that protein kinases are located in the mitochondria of certain tissues and species. These include pyruvate dyhydrogenase kinase, branched-chain alpha-ketoacid dehydrogenase kinase, protein kinase A, protein kinase Cdelta, stress-activated kinase and A-Raf as well as unidentified kinases. Recent evidence suggests that mitochondrial protein kinases may be involved in physiological processes such as apoptosis and steroidogenesis. Additionally, the novel finding of low-molecular-weight GTP-binding proteins in mitochondria suggests the possibility that these may interact with mitochondrial protein kinases to regulate the activity of mitochondrial effector proteins. The fact that there are components of cellular regulatory systems in mitochondria indicates the exciting possibility of undiscovered systems regulating mitochondrial physiology.

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“…In other words, when these structures are aligned to the PDK2 model with TOP3D (17), they position a positive charge less than 2 Å from the ⑀-nitrogen of Lys-246 in our structure. Given this catalytic similarity of the ATP-binding domain of PDK2 with the family of ATPases, it is not surprising that PDK2 has been found to also possess an ATPase activity, representing 1-3% of total kinase activity (33).…”

Section: (Atp Lid)mentioning

confidence: 99%

Structure of Pyruvate Dehydrogenase Kinase

Steussy

Popov

Bowker-Kinley

et al. 2001

Journal of Biological Chemistry

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The structure of mitochondrial pyruvate dehydrogenase kinase isozyme 2 is of interest because it represents a family of serine-specific protein kinases that lack sequence similarity with all other eukaryotic protein kinases. Similarity exists instead with key motifs of prokaryotic histidine protein kinases and a family of eukaryotic ATPases. The 2.5-Å crystal structure reported here reveals that pyruvate dehydrogenase kinase isozyme 2 has two domains of about the same size. The N-terminal half is dominated by a bundle of four amphipathic ␣-helices, whereas the C-terminal half is folded into an ␣/␤ sandwich that contains the nucleotide-binding site. Analysis of the structure reveals this C-terminal domain to be very similar to the nucleotidebinding domain of bacterial histidine kinases, but the catalytic mechanism appears similar to that of the eukaryotic serine kinases and ATPases.

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An Essential Role of Glu-243 and His-239 in the Phosphotransfer Reaction Catalyzed by Pyruvate Dehydrogenase Kinase

Cited by 23 publications

References 36 publications

Autophosphorylation Activity of the Arabidopsis Ethylene Receptor Multigene Family

Autophosphorylation Activity of the Arabidopsis Ethylene Receptor Multigene Family

Evidence of undiscovered cell regulatory mechanisms: phosphoproteins and protein kinases in mitochondria

Structure of Pyruvate Dehydrogenase Kinase

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