The peripheral subunit-binding domain of the dihydrolipoyl acetyltransferase component of the pyruvate dehydrogenase complex of Bacillus stearothermophilus: preparation and characterization of its binding to the dihydrolipoyl dehydrogenase component.

  • Hipps D
  • Packman L
  • Allen M
 et al. 
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Abstract

The peripheral subunit-binding domain of the dihydrolipoyl acetyltransferase polypeptide chain of the pyruvate dehydrogenase multienzyme complex of Bacillus stearothermophilus was released by limited proteolysis from a di-domain (lipoyl domain plus binding domain) encoded by a subgene over-expressed in Escherichia coli. The domain was characterized by N-terminal sequence analysis, electrospray m.s. and c.d. spectroscopy. It was found to be identical in all respects to a chemically synthesized peptide of the same sequence. The association of the di-domain and binding domain (both natural and synthetic) with dihydrolipoyl dehydrogenase was analysed in detail and a tight binding was demonstrated. As judged by several different techniques, it was found that only one peripheral subunit-binding domain is bound to one dimer of dihydrolipoyl dehydrogenase, implying that the association is highly anti-cooperative.

Author-supplied keywords

  • Amino Acid Sequence
  • Binding Sites
  • Circular Dichroism
  • Dihydrolipoamide Dehydrogenase
  • Dihydrolipoamide Dehydrogenase: chemistry
  • Dihydrolipoamide Dehydrogenase: metabolism
  • Geobacillus stearothermophilus
  • Geobacillus stearothermophilus: enzymology
  • Magnetic Resonance Spectroscopy
  • Mass Spectrometry
  • Molecular Sequence Data
  • Peptide Fragments
  • Peptide Fragments: chemistry
  • Peptide Fragments: metabolism
  • Pyruvate Dehydrogenase Complex
  • Pyruvate Dehydrogenase Complex: chemistry
  • Pyruvate Dehydrogenase Complex: metabolism
  • Recombinant Proteins
  • Recombinant Proteins: chemistry
  • Recombinant Proteins: metabolism
  • Sequence Analysis
  • Trypsin
  • Trypsin: metabolism

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Authors

  • D S Hipps

  • L C Packman

  • M D Allen

  • C Fuller

  • K Sakaguchi

  • E Appella

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