title: Expression and lipoylation in Escherichia coli of the inner lipoyl domain of the E2 component of the human pyruvate dehydrogenase complex.
abstract: The dihydrolipoamide acetyltransferase subunit (E2p) of mammalian pyruvate dehydrogenase complex has two highly conserved lipoyl domains each modified with a lipoyl cofactor bound in amide linkage to a specific lysine residue. A sub-gene encoding the inner lipoyl domain of human E2p has been over-expressed in Escherichia coli. Two forms of the domain have been purified, corresponding to lipoylated and non-lipoylated species. The apo-domain can be lipoylated in vitro with partially purified E. coli lipoate protein ligase, and the lipoylated domain can be reductively acetylated by human E1p (pyruvate dehydrogenase). Availability of the two forms will now allow detailed biochemical and structural studies of the human lipoyl domains.
authors.affiliation: Department of Biochemistry and Genetics, Medical School, University of Newcastle upon Tyne, U.K.
authors.forename: J
authors.initials: J
authors.lastname: Quinn
authors.forename: A G
authors.initials: AG
authors.lastname: Diamond
authors.forename: A K
authors.initials: AK
authors.lastname: Masters
authors.forename: D E
authors.initials: DE
authors.lastname: Brookfield
authors.forename: N G
authors.initials: NG
authors.lastname: Wallis
authors.forename: S J
authors.initials: SJ
authors.lastname: Yeaman
mesh_terms: D000123:Acetyltransferases / Q000737:chemistry / Q000235:genetics*; D000595:Amino Acid Sequence; D001483:Base Sequence; D002852:Chromatography, Ion Exchange; D003001:Cloning, Molecular; D004274:DNA, Recombinant; D051047:Dihydrolipoyllysine-Residue Acetyltransferase; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D005982:Glutathione Transferase / Q000737:chemistry; D006801:Humans; D008969:Molecular Sequence Data; D010453:Peptide Synthases / Q000378:metabolism; D010957:Plasmids; D011768:Pyruvate Dehydrogenase Complex / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics
chemical_list: D004274:DNA, Recombinant; D011768:Pyruvate Dehydrogenase Complex; D011993:Recombinant Fusion Proteins; D000123:Acetyltransferases; D051047:Dihydrolipoyllysine-Residue Acetyltransferase; D005982:Glutathione Transferase; D010453:Peptide Synthases; C072322:lipoate-protein ligase