Structural and functional peculiarities of alpha-ketoglutarate dehydrogenase with non-interacting active sites.
The properties of alpha-ketoglutarate dehydrogenase with non-interacting active sites were investigated. The substrate and coenzyme saturation curves are found to be hyperbolic, which is consistent with the absence of cooperativity between the active sites of the enzyme. The peculiarities of KGD of this form, determining its functional properties, were revealed. Thus, 6 cysteine residues of the enzyme possess different properties in comparison with the form of the enzyme with interacting active sites. 3 Sulfhydryl groups of the "non-cooperative" enzyme form were rapidly oxidized in the process of the enzyme isolation and storage; thereafter they could not be reduced by dithiols. Three other cysteine residues are probably involved in the formation of disulfide bonds. Two of them are supposed to form intramolecular disulfide, whereas the third one is thought to be modified by some low molecular weight disulfide. The reduction of these sulfhydryl groups by dithiols is shown to be accompanied by the appearance of the kinetic cooperativity with respect to the substrate. It is suggested that the thiol/disulfide exchange in vivo can regulate a reversible conversion of the "non-cooperative" KGD form into one with interacting sites.
Bunik
V I
VI
Department of Biochemistry, Moscow State University, USSR.
Buneeva
O A
OA
Lvova
N B
NB
Gomazkova
V S
VS
D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D010856:Columbidae; D003545:Cysteine / Q000032:analysis; D004220:Disulfides / Q000378:metabolism; D004795:Enzyme Stability; D007655:Ketoglutarate Dehydrogenase Complex / Q000302:isolation & purification*; D007658:Ketone Oxidoreductases / Q000302:isolation & purification*; D007700:Kinetics; D009132:Muscles / Q000201:enzymology*; D010084:Oxidation-Reduction; D013329:Structure-Activity Relationship; D013438:Sulfhydryl Compounds / Q000032:analysis / Q000378:metabolism
D004220:Disulfides; D013438:Sulfhydryl Compounds; D007658:Ketone Oxidoreductases; D007655:Ketoglutarate Dehydrogenase Complex; D003545:Cysteine