The carboxy-terminal tail of pyruvate dehydrogenase kinase 2 is required for the kinase activity

Biochemistry. 2005 Oct 18;44(41):13573-82. doi: 10.1021/bi0505868.

Abstract

Pyruvate dehydrogenase kinase 2 (PDK2) is a prototypical mitochondrial protein kinase that regulates the activity of the pyruvate dehydrogenase complex. Recent structural studies have established that PDK2 consists of a catalytic core built of the B and K domains and the relatively long amino and carboxyl tails of unknown function. Here, we show that the carboxy-terminal truncation variants of PDK2 display a greatly diminished capacity for phosphorylation of holo-PDC. This effect is due largely to the inability of the transacetylase component of PDC to promote the phosphorylation reaction catalyzed by the truncated PDK2 variants. Furthermore, the truncated forms of PDK2 bind poorly to the lipoyl-bearing domain(s) provided by the transacetylase component. Taken together, these data strongly suggest that the carboxyl tails of PDK isozymes contribute to the lipoyl-bearing domain-binding site of the kinase molecule. We also show that the carboxyl tails derived from isozymes PDK1, PDK3, and PDK4 are capable of supporting the kinase activity of the kinase core derived from PDK2 as well as binding of the respective PDK2 chimeras to the lipoyl-bearing domain. Furthermore, the chimera carrying the carboxyl tail of PDK3 displays a stronger response to the addition of the transacetylase component along with a better binding to the lipoyl-bearing domain, suggesting that, at least in part, the differences in the amino acid sequences of the carboxyl tails account for the differences between PDK isozymes.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Adenosine Diphosphate / metabolism
  • Amino Acid Sequence
  • Animals
  • Calorimetry
  • Electrophoresis, Polyacrylamide Gel
  • Molecular Sequence Data
  • Protein Kinases / chemistry*
  • Protein Kinases / metabolism*
  • Protein Structure, Tertiary
  • Protein-Serine-Threonine Kinases
  • Pyruvate Dehydrogenase (Acetyl-Transferring) Kinase
  • Pyruvate Dehydrogenase Complex / metabolism
  • Rats
  • Sequence Alignment
  • Thermodynamics

Substances

  • Pdk1 protein, rat
  • Pdk2 protein, rat
  • Pdk3 protein, rat
  • Pdk4 protein, rat
  • Pyruvate Dehydrogenase (Acetyl-Transferring) Kinase
  • Pyruvate Dehydrogenase Complex
  • Adenosine Diphosphate
  • Protein Kinases
  • Protein-Serine-Threonine Kinases