title: 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.
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.
authors.affiliation: Department of Biochemistry, University of Cambridge, U.K.
authors.forename: D S
authors.initials: DS
authors.lastname: Hipps
authors.forename: L C
authors.initials: LC
authors.lastname: Packman
authors.forename: M D
authors.initials: MD
authors.lastname: Allen
authors.forename: C
authors.initials: C
authors.lastname: Fuller
authors.forename: K
authors.initials: K
authors.lastname: Sakaguchi
authors.forename: E
authors.initials: E
authors.lastname: Appella
authors.forename: R N
authors.initials: RN
authors.lastname: Perham
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D002942:Circular Dichroism; D008058:Dihydrolipoamide Dehydrogenase / Q000737:chemistry / Q000378:metabolism*; D001411:Geobacillus stearothermophilus / Q000201:enzymology*; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D011768:Pyruvate Dehydrogenase Complex / Q000737:chemistry / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D017421:Sequence Analysis; D014357:Trypsin / Q000378:metabolism
chemical_list: D010446:Peptide Fragments; D011768:Pyruvate Dehydrogenase Complex; D011994:Recombinant Proteins; D008058:Dihydrolipoamide Dehydrogenase; D014357:Trypsin