Inhibition of the bovine branched-chain 2-oxo acid dehydrogenase complex and its kinase by arylidenepyruvates.
A novel class of inhibitors for the branched-chain 2-oxo acid dehydrogenase (BCOAD) complex has been synthesized and studied. The sodium salts of arylidenepyruvates: e.g., furfurylidenepyruvate (compound I), 4-(3-thienyl)-2-oxo-3-butenoate (compound II), cinnamalpyruvate (compound III) and 4-(2-thienyl)-2-oxo-3-butenoate (compound IV) inhibit the overall and kinase reactions of the BCOAD complex from bovine liver. Inhibitions of the overall reaction occur at the decarboxylase (E1) step as determined by a spectrophotometric assay with 2,6-dichlorophenolindophenol as an electron acceptor. Inhibition of the E1 reaction by compound I (Ki = 0.5 microM) is competitive, whereas inhibitions by compounds II (Ki = 150 microM) and III (Ki = 500 microM) are non-competitive with respect to the substrate 2-oxoisovalerate. The Km value for 2-oxoisovalerate is 6.7 microM as measured by the E1 assay. Inhibition of the E1 step by compounds I, II and III are reversible at low inhibitor concentrations based on the Michaelis-Menten kinetics observed. By comparison, compound I does not significantly inhibit pyruvate and 2-oxoglutarate dehydrogenase complexes. The arylidenepyruvates (compounds I, II and IV) inhibit the BCOAD kinase reaction in a manner similar to the substrate 2-oxo acids. The inhibition of the kinase reaction by compound I is non-competitive with respect to ATP, with an apparent Ki value of 4.5 mM. The results suggest that arylidenepyruvates may be useful probes for elucidating the reaction mechanisms of the BCOAD complex and its kinase.
Lau
K S
KS
Department of Biochemistry, University of Texas Southwestern Medical Center, Dallas 75235-9038.
Cooper
A J
AJ
Chuang
D T
DT
D042942:3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide); D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D002417:Cattle; D055598:Chemical Phenomena; D002621:Chemistry; D002854:Chromatography, Paper; D004789:Enzyme Activation / Q000187:drug effects; D007192:Indenes / Q000494:pharmacology*; D007658:Ketone Oxidoreductases / Q000037:antagonists & inhibitors*; D007700:Kinetics; D009097:Multienzyme Complexes / Q000037:antagonists & inhibitors*; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases; D011773:Pyruvates / Q000494:pharmacology*
D007192:Indenes; D009097:Multienzyme Complexes; D047428:Protein Kinase Inhibitors; D011773:Pyruvates; D000255:Adenosine Triphosphate; D007658:Ketone Oxidoreductases; D042942:3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide); D011494:Protein Kinases; C035627:(3-methyl-2-oxobutanoate dehydrogenase (lipoamide)) kinase