A novel branched-chain amino acid metabolon. Protein-protein interactions in a supramolecular complex.
The catabolic pathways of branched-chain amino acids have two common steps. The first step is deamination catalyzed by the vitamin B(6)-dependent branched-chain aminotransferase isozymes (BCATs) to produce branched-chain alpha-keto acids (BCKAs). The second step is oxidative decarboxylation of the BCKAs mediated by the branched-chain alpha-keto acid dehydrogenase enzyme complex (BCKD complex). The BCKD complex is organized around a cubic core consisting of 24 lipoate-bearing dihydrolipoyl transacylase (E2) subunits, associated with the branched-chain alpha-keto acid decarboxylase/dehydrogenase (E1), dihydrolipoamide dehydrogenase (E3), BCKD kinase, and BCKD phosphatase. In this study, we provide evidence that human mitochondrial BCAT (hBCATm) associates with the E1 decarboxylase component of the rat or human BCKD complex with a K(D) of 2.8 microM. NADH dissociates the complex. The E2 and E3 components do not interact with hBCATm. In the presence of hBCATm, k(cat) values for E1-catalyzed decarboxylation of the BCKAs are enhanced 12-fold. Mutations of hBCATm proteins in the catalytically important CXXC center or E1 proteins in the phosphorylation loop residues prevent complex formation, indicating that these regions are important for the interaction between hBCATm and E1. Our results provide evidence for substrate channeling between hBCATm and BCKD complex and formation of a metabolic unit (termed branched-chain amino acid metabolon) that can be influenced by the redox state in mitochondria.
Islam
Mohammad Mainul
MM
Department of Biochemistry, Wake Forest University School of Medicine, Winston-Salem, North Carolina 27157, USA.
Wallin
Reidar
R
Wynn
R Max
RM
Conway
Myra
M
Fujii
Hisao
H
Mobley
James A
JA
Chuang
David T
DT
Hutson
Susan M
SM
D000597:Amino Acids, Branched-Chain / Q000737:chemistry*; D000818:Animals; D002384:Catalysis; D006801:Humans; D007700:Kinetics; D008297:Male; D008928:Mitochondria / Q000378:metabolism*; D008930:Mitochondria, Liver / Q000378:metabolism; D009243:NAD / Q000737:chemistry; D025941:Protein Interaction Mapping; D017434:Protein Structure, Tertiary; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000737:chemistry; D000637:Transaminases / Q000737:chemistry
D000597:Amino Acids, Branched-Chain; D011994:Recombinant Proteins; D009243:NAD; D000637:Transaminases