Isolation of glyoxalase II from two different compartments of rat liver mitochondria. Kinetic and immunochemical characterization of the enzymes.
Two separate pools of glyoxalase II were demonstrated in rat liver mitochondria, one in the intermembrane space and the other in the matrix. The enzyme was purified from both sources by affinity chromatography on S-(carbobenzoxy)glutathione-Affi-Gel 40. From both crude and purified preparations polyacrylamide gel-electrophoresis resolved multiple forms of glyoxalase II, two from the intermembrane space and five from the matrix. Among the thioesters of glutathione tested as substrates, S-D-lactoylglutathione was hydrolyzed most efficiently by the enzymes from both sources. Significant differences were observed in the specificities between the intermembrane space and matrix enzymes with S-acetoacetylglutathione, S-acetylglutathione, S-propionylglutathione and S-succinylglutathione as substrates. Pure glyoxalase II from rat liver cytosol was chemically polymerized and used as antigen. Antibodies were raised in rabbits and the antiserum was used for comparison of the two purified mitochondrial enzymes with cytosolic glyoxalase II by immunoblotting. The enzyme purified from the intermembrane space cross-reacted with the antiserum, but the matrix glyoxalase II did not. The results give evidence for the presence in rat liver mitochondria of two species of glyoxalase II with differing characteristics. Only the enzyme from the intermembrane space appears to resemble the cytosolic glyoxalase II forms.
Talesa
V
V
Department of Medical Chemistry, University of Helsinki, Finland.
Uotila
L
L
Koivusalo
M
M
Principato
G
G
Giovannini
E
E
Rosi
G
G
D000818:Animals; D002846:Chromatography, Affinity; D004589:Electrophoresis, Disc; D005260:Female; D015151:Immunoblotting; D007425:Intracellular Membranes / Q000201:enzymology*; D007527:Isoenzymes / Q000302:isolation & purification* / Q000378:metabolism; D007700:Kinetics; D008297:Male; D008930:Mitochondria, Liver / Q000201:enzymology*; D008970:Molecular Weight; D051381:Rats; D013367:Submitochondrial Particles / Q000201:enzymology*; D013379:Substrate Specificity; D013869:Thiolester Hydrolases / Q000302:isolation & purification* / Q000378:metabolism; D000099301:Hydroxyacylglutathione Hydrolase
D007527:Isoenzymes; D013869:Thiolester Hydrolases; D000099301:Hydroxyacylglutathione Hydrolase