Purification and characterization of human liver branched-chain α-keto acid dehydrogenase complex

Ôno, Kazuo; Hakozaki, Michiyuki; Nishimaki, H.; Kochi, Hideo

doi:10.1016/0885-4505(87)90019-3

Cited by 17 publications

(7 citation statements)

References 20 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…E3 remained in the two top fractions regardless of the presence or absence of E1 and/or E2. In this regard, it is known that E3 is easily released from BCKADH [14,15]. When E3 was not added, however, the E1 recovered in fractions 3, 4 and the pellet fraction was significantly reduced (data not shown), probably due to inappropriate interaction between E1 and E2 in the absence of E3.…”

Section: Resultsmentioning

confidence: 92%

“…Figure 2B shows the nucleotide sequence and the predicted amino‐acid sequence of chicken E2. Chicken E2 appears to conserve three functional domains: the lipoyl‐bearing domain (residues 1–72), the E1/E3‐binding domain (residues 109–152) and the inner‐core domain (residues 186–432) as in mammalian E2 [4–20,32]. The nucleotide sequence of chicken E1β cDNA (1543 bp in length) is not shown, as the deduced amino‐acid sequence of mature chicken E1β was highly homologous to that of mammalian E1β (see below) and not characterized by a chicken‐specific sequence.…”

Section: Resultsmentioning

confidence: 99%

“…Rat liver was prepared from males of the Wistar strain. Purified human E1 and E2 were the preparations employed in our previous study [15]. Leupeptin and trypsin inhibitor were purchased from Protein Institute (Osaka, Japan).…”

Section: Methodsmentioning

confidence: 99%

“…On the other hand, the E3 component, a homodimer [11], is also a catalytic component of the pyruvate dehydrogenase complex, the 2‐oxoglutarate dehydrogenase complex and the glycine cleavage system [12]. Studies with purified BCKADH components [13–15] and molecular cloning of cDNA for E1α[16–18], E1β[19–22] and E2 [7,23,24] of various mammals have revealed that the amino‐acid sequences of respective polypeptides are highly homologous among mammalian species.…”

mentioning

confidence: 99%

See 3 more Smart Citations

cDNA cloning of the chicken branched‐chain α‐keto acid dehydrogenase complex

Ôno¹,

Hakozaki²,

Suzuki³

et al. 2001

European Journal of Biochemistry

View full text Add to dashboard Cite

Branched-chain a-keto acid dehydrogenase complex is a macromolecule comprising three catalytic components: a dehydrogenase (E1) with a 2 b 2 structure, an acyltransferase (E2) and a dihydrolipoamide dehydrogenase (E3). In the mammalian complex, the E2 component with 24 identical subunits forms a structural core, to which multiple copies of E1 and E3 bind noncovalently. We isolated cDNA clones encoding E1a, E1b and E2 subunits from a chicken-liver cDNA library and performed nucleotide sequencing. Amino-acid sequences deduced from the nucleotide sequences revealed that chicken E1a and E1b chains had substantially homologous sequences with the corresponding mammalian polypeptides, except for the N-terminus. Chicken E2 conserved three functional domains, a lipoylbearing domain, an E1/E3 binding domain and an innercore domain, but contrasted strongly with mammalian E2 in respect of containing 11 additional residues in two interdomain linkers: nine sequential residues in one linker and two residues in the other. Replacement of many residues was also observed in the chicken linkers. When E2 activity for catalyzing the overall reaction was measured by activity reconstitution in combination with E1 and E3, chicken E2 was markedly less effective than mammalian E2. The capability of chicken E2 for binding E1 was also reduced when determined by the binding assay using sucrose density gradient centrifugation. Chicken E1 was functionally as well as structurally indistinguishable from mammalian E1. Thus the reduced catalytic activity of chicken E2 must arise from its reduced E1-binding capacity, which results from the characteristic structure of interdomain linkers in chicken E2.

show abstract

Section: Resultsmentioning

confidence: 92%

Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

See 2 more Smart Citations

cDNA cloning of the chicken branched‐chain α‐keto acid dehydrogenase complex

Ôno¹,

Hakozaki²,

Suzuki³

et al. 2001

European Journal of Biochemistry

View full text Add to dashboard Cite

show abstract

“…The BCOADC activity was reconstituted in vitro by mixing the purified E1, E2, and E3 components as described previously [24]. Before assay, 1.25 μ g of E1 was incubated for 3 min on ice with sera at a 200-fold dilution.…”

Section: Methodsmentioning

confidence: 99%

Characterization of Autoantibodies against the E1 Subunit of Branched-Chain 2-Oxoacid Dehydrogenase in Patients with Primary Biliary Cirrhosis

Mori

Ohira

Kuroda

et al. 2012

International Journal of Hepatology

View full text Add to dashboard Cite

Primary biliary cirrhosis (PBC) is characterized by antimitochondrial antibodies (AMAs) that react with the lipoyl-containing E2 subunits of 2-oxoacid dehydrogenase complexes such as BCOADC and PDC. The lipoyl domains of E2 contain the major epitopes essential for immunopathology. However, the non-lipoyl-containing E1 subunits are also frequently targeted. Since anti-E1 antibodies always appear in combination with anti-E2 antibodies, the mechanisms underlying the autoimmunity against E1 may be linked to, but distinct from, those against E2. Here, we demonstrate that intermolecular and intramolecular determinant spreading underlies the autoimmunity against E1. We performed characterizations and epitope mapping for anti-BCOADC-E1 antibodies from both the intermolecular and intramolecular points of view. The antibody reactivities form a cluster against the BCOADC complex that is distinct from that against the PDC complex, and the anti-BCOADC-E1 antibodies arise as part of the cluster against the BCOADC complex. Multiple epitopes are present on the surface of the BCOADC-E1 molecule, and the major epitope overlaps with the active center. Sera with anti-BCOADC-E1 antibodies strongly inhibited the enzyme activity. These findings suggest that the E1 subunit as part of the native BCOADC complex is an immunogen, and that determinant spreading is involved in the pathogenesis of AMA production.

show abstract

Molecular defects of the branched-chain α-keto acid dehydrogenase complex: Maple syrup urine disease due to mutations of the E1α or E1β subunit gene

Indo

Matsuda

1996

Alpha-Keto Acid Dehydrogenase Complexes

View full text Add to dashboard Cite

Purification and characterization of human liver branched-chain α-keto acid dehydrogenase complex

Cited by 17 publications

References 20 publications

cDNA cloning of the chicken branched‐chain α‐keto acid dehydrogenase complex

cDNA cloning of the chicken branched‐chain α‐keto acid dehydrogenase complex

Characterization of Autoantibodies against the E1 Subunit of Branched-Chain 2-Oxoacid Dehydrogenase in Patients with Primary Biliary Cirrhosis

Molecular defects of the branched-chain α-keto acid dehydrogenase complex: Maple syrup urine disease due to mutations of the E1α or E1β subunit gene

Contact Info

Product

Resources

About