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 appl ied to study the 2-oxoacid:lipoate oxidoreductase activity of 2-oxoaci d dehydrogenase complexes, The enzymatic and non-enzymatic mechanisms of the transfer of reducing equivalents from the complexes to free lip oic acid (alpha-lipoic acid, 6,8-thiooctic acid) were distinguished us ing the high stereoselectivity of the complex enzymes to the R-enantio mer of lipoate, Unlike these enzymes, thioredoxin from E, coli exibite d no stereoselectivity upon reduction,vith chemically obtained dihydro lipoate, However, coupled to the dihydrolipoate production by the dehy drogenase complexes, the process was essentially sensitive both to the enantiomer used and the dihydrolipoyl dehydrogenase activity of the c omplexes, These results indicated the involvement of the third complex component, dihydrolipoyl dehydrogenase, in the 2-oxoacid-dependent di hydrolipoate formation, The implication of the investigated reaction f or a connection between thioredoxin and the 2-oxoacid dehydrogenase co mplexes in the mitochondrial metabolism are discussed.