Evidence that isoenzymes i and 2 of pyruvate dehydrogenase kinase (PDK) can form heterodimers in vitro

Kirill M. Popov, Robert Harris

Research output: Contribution to journalArticle

Abstract

Three isoenzymes of PDK that different in terms of their activities and regulatory properties have been identified in mammalian tissues. It appears that each tissue tested thus far expresses its own subset of isoenzymes, suggesting that overall kinase activity of any particular tissue reflects the isoenzymic composition of PDK. It is generally believed that physically the mammalian PDK is a dimer. However, whether the different isoenzymes of PDK can interact with each other has not been established yet. Here we report evidence that isoenzymes f (PDKI ) and 2 (PDK2) of rat PDK can form heterodimers in vitro. The sequence of PDKI was modified to carry T7-Tag at its amino terminus in order to monitor its expression with anti-T7-Tag monoclonal antibodies. The sequence of PDK2 was modified to carry T7-Tag as well as HisTag to facilitate the purification by metal chelate chromatography. Resulting cDNAs were expressed in Escherichia coli under control of bacteriophage T7 promoter. Analysis of the recombinant kinase purified on His-Bind resin revealed that isoenzymes I and 2 preferentially form heterodimers when expressed simultaneously. Heterodimers, in contrast to PDKI and PDK2 homodimers, demonstrated diminished response to NADH and acetyl-CoA, factors considered to be involved in regulation of kinase activity in vivo. We propose, therefore, that the response of PDK to the different metabolic signals may be modulated by the cross-dimerization of PDK isoenzymes.

Original languageEnglish
JournalFASEB Journal
Volume10
Issue number6
StatePublished - 1996

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pyruvate dehydrogenase (acetyl-transferring) kinase
Isoenzymes
isozymes
phosphotransferases (kinases)
Phosphotransferases
Tissue
Bacteriophage T7
Acetyl Coenzyme A
Bacteriophages
dimerization
Dimerization
affinity chromatography
Chromatography
In Vitro Techniques
Dimers
bacteriophages
NAD
Escherichia coli
resins
Purification

ASJC Scopus subject areas

  • Agricultural and Biological Sciences (miscellaneous)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Biochemistry
  • Cell Biology

Cite this

Evidence that isoenzymes i and 2 of pyruvate dehydrogenase kinase (PDK) can form heterodimers in vitro. / Popov, Kirill M.; Harris, Robert.

In: FASEB Journal, Vol. 10, No. 6, 1996.

Research output: Contribution to journalArticle

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