title: Requirements for the adaptor protein role of dihydrolipoyl acetyltransferase in the up-regulated function of the pyruvate dehydrogenase kinase and pyruvate dehydrogenase phosphatase.
abstract: The dihydrolipoyl acetyltransferase (E2 component) is a 60-mer assembled via its COOH-terminal domain with exterior E1-binding domain and two lipoyl domains (L2 then L1) sequentially connected by mobile linker regions. E2 facilitates markedly enhanced function of the pyruvate dehydrogenase kinase (PDK) and pyruvate dehydrogenase phosphatase (PDP). Human E2 structures were prepared with only one lipoyl domain (L1 or L2) or with alanines substituted at the sites of lipoylation (Lys-46 in L1 or Lys-173 in L2). The L2 domain and its lipoyl group were shown to be essential for markedly enhanced PDP function and were required for greatly up-regulated PDK function. The complete absence of the L1 domain reduced the enhancements of both of these activities but not the maximal effector-stimulated PDK activity through acetylation of L2. With nonlipoylated L2 present, lipoylated L1 supported a lesser enhancement in PDK function with significant stimulation upon acetylation of L1. Prevention of L1 lipoylation in K46AE2 removed this competitive L1 role and enhanced L2-facilitated PDK activity beyond that of native E2 when PDK activity was measured in the absence or in the presence of stimulatory effectors. Thus, the E2-L2 domain has a paramount role in facilitating enhanced PDK and PDP function but inclusion of E2-L1 domain, even in a noninteracting (nonlipoylated) form, contributes to the marked elevation of these activities.
authors.affiliation: Department of Biochemistry, Kansas State University, Manhattan, Kansas 66506, USA.
authors.forename: D
authors.initials: D
authors.lastname: Yang
authors.forename: X
authors.initials: X
authors.lastname: Gong
authors.forename: A
authors.initials: A
authors.lastname: Yakhnin
authors.forename: T E
authors.initials: TE
authors.lastname: Roche
mesh_terms: D000123:Acetyltransferases / Q000737:chemistry*; D000818:Animals; D002417:Cattle; D051047:Dihydrolipoyllysine-Residue Acetyltransferase; D004789:Enzyme Activation / Q000502:physiology; D015971:Gene Expression Regulation, Enzymologic / Q000235:genetics; D006801:Humans; D011485:Protein Binding; D011494:Protein Kinases / Q000502:physiology*; D017346:Protein Serine-Threonine Kinases; D011769:Pyruvate Dehydrogenase (Lipoamide)-Phosphatase / Q000502:physiology*; D000081382:Pyruvate Dehydrogenase Acetyl-Transferring Kinase; D011768:Pyruvate Dehydrogenase Complex / Q000502:physiology; D011994:Recombinant Proteins / Q000737:chemistry; D008063:Thioctic Acid / Q000378:metabolism; D015854:Up-Regulation / Q000502:physiology*
chemical_list: D000081382:Pyruvate Dehydrogenase Acetyl-Transferring Kinase; D011768:Pyruvate Dehydrogenase Complex; D011994:Recombinant Proteins; D008063:Thioctic Acid; D000123:Acetyltransferases; D051047:Dihydrolipoyllysine-Residue Acetyltransferase; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D011769:Pyruvate Dehydrogenase (Lipoamide)-Phosphatase