Interactions between 4-aminobutyrate aminotransferase and succinic semialdehyde dehydrogenase, two mitochondrial enzymes.
Physical interactions between the enzymes involved in the catabolism of the neurotransmitter 4-aminobutyrate were detected by means of affinity chromatography and fluorescence techniques. By immobilizing one enzyme (4-aminobutyrate aminotransferase) indirectly through antibodies bound to protein A-Sepharose, it was possible to demonstrate that succinic semialdehyde dehydrogenase interacts with the aminotransferase at neutral pH and ionic strength values higher than 0.2. Increasing the ionic strength of the medium results in dissociation of the "enzyme cluster." Binding of succinic semialdehyde dehydrogenase to the aminotransferase tagged with a fluorescent probe was detected by polarization of fluorescence measurements at neutral pH. Upon saturation of the aminotransferase with succinic semialdehyde dehydrogenase, the polarization of fluorescence increases from 0.13 to 0.21. The results are consistent with a model in which one molecule of succinic semialdehyde dehydrogenase is bound to one molecule of 4-aminobutyrate aminotransferase with an equilibrium dissociation constant of 0.1 microM. Since the concentrations of both enzymes in the mitochondrial matrix have been estimated to be around 2 microM, the results obtained with the purified mitochondrial enzymes strongly suggest that the aminotransferase is saturated with succinic semialdehyde dehydrogenase to form a stable enzymatic complex under in vivo conditions.
D000612:4-Aminobutyrate Transaminase / Q000378:metabolism*; D000818:Animals; D001921:Brain / Q000201:enzymology*; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D004800:Enzymes, Immobilized / Q000378:metabolism; D005454:Fluorescence Polarization; D006863:Hydrogen-Ion Concentration; D006884:Hydroxybutyrate Dehydrogenase / Q000378:metabolism*; D007700:Kinetics; D008433:Mathematics; D008928:Mitochondria / Q000201:enzymology*; D009994:Osmolar Concentration; D050644:Succinate-Semialdehyde Dehydrogenase; D013552:Swine
D004800:Enzymes, Immobilized; D006884:Hydroxybutyrate Dehydrogenase; C019416:4-hydroxybutyrate dehydrogenase; C493643:ALDH5A1 protein, human; D050644:Succinate-Semialdehyde Dehydrogenase; D000612:4-Aminobutyrate Transaminase