Using lipoate enantiomers and thioredoxin to study the mechanism of the 2-oxoacid-dependent dihydrolipoate production by the 2-oxoacid dehydrogenase complexes.
The thioredoxin-catalyzed insulin reduction by dihydrolipoate was applied to study the 2-oxoacid: lipoate oxidoreductase activity of 2-oxoacid dehydrogenase complexes. The enzymatic and non-enzymatic mechanisms of the transfer of reducing equivalents from the complexes to free lipoic acid (alpha-lipoic acid, 6,8-thiooctic acid) were distinguished using the high stereoselectivity of the complex enzymes to the R-enantiomer of lipoate. Unlike these enzymes, thioredoxin from E. coli exhibited no stereoselectivity upon reduction with chemically obtained dihydrolipoate. However, coupled to the dihydrolipoate production by the dehydrogenase complexes, the process was essentially sensitive both to the enantiomer used and the dihydrolipoyl dehydrogenase activity of the complexes. These results indicated the involvement of the third complex component, dihydrolipoyl dehydrogenase, in the 2-oxoacid-dependent dihydrolipoate formation. The implication of the investigated reaction for a connection between thioredoxin and the 2-oxoacid dehydrogenase complexes in the mitochondrial metabolism are discussed.
Bunik
V
V
A.N. Belozersky Institute of Physico-Chemical Biology, Moscow State University, Russian Federation.
Shoubnikova
A
A
Loeffelhardt
S
S
Bisswanger
H
H
Borbe
H O
HO
Follmann
H
H
D042942:3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide); D001713:Biphenyl Compounds / Q000494:pharmacology; D011232:Chemical Precipitation; D004926:Escherichia coli / Q000201:enzymology; D005182:Flavin-Adenine Dinucleotide / Q000378:metabolism / Q000494:pharmacology; D007328:Insulin / Q000378:metabolism; D007655:Ketoglutarate Dehydrogenase Complex / Q000378:metabolism; D007656:Ketoglutaric Acids / Q000494:pharmacology*; D007658:Ketone Oxidoreductases / Q000378:metabolism*; D009097:Multienzyme Complexes / Q000378:metabolism*; D009243:NAD / Q000494:pharmacology; D009861:Onium Compounds / Q000494:pharmacology; D011768:Pyruvate Dehydrogenase Complex / Q000378:metabolism; D013237:Stereoisomerism; D008063:Thioctic Acid / Q000031:analogs & derivatives* / Q000378:metabolism*; D013879:Thioredoxins / Q000378:metabolism*
D001713:Biphenyl Compounds; D007328:Insulin; D007656:Ketoglutaric Acids; D009097:Multienzyme Complexes; D009861:Onium Compounds; D011768:Pyruvate Dehydrogenase Complex; D009243:NAD; C031291:diphenyliodonium; D005182:Flavin-Adenine Dinucleotide; D013879:Thioredoxins; D008063:Thioctic Acid; C015144:dihydrolipoic acid; D007658:Ketone Oxidoreductases; D007655:Ketoglutarate Dehydrogenase Complex; D042942:3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)