Glyoxalase 2 deficiency in the erythrocytes of a horse: 1H NMR studies of enzyme kinetics and transport of S-lactoylglutathione.
In mammalian red blood cells the metabolism of methylglyoxal, and some alpha-ketoaldehydes, takes place via two, generally, highly active enzymes, glyoxalase 1 and 2. The 1H NMR spin-echo spectra of horse erythrocytes, and the various reactants in the glyoxalase system, were characterized as a prelude to obtaining series of spectra in time courses of methylglyoxal metabolism. We characterized the kinetics of the enzyme system in red cells from a normal horse and also from one which had very low activity of glyoxylase 2. The kinetics of the reaction scheme, with methylglyoxal as the starting substrate, were obtained from 1H NMR spectra and analyzed with a computer model of the scheme. The most salient feature of the normal system was the very high feed-forward inhibition (KiHTA = 0.1 microM) of glyoxalase 2 by the hemithioacetal which is the substrate of glyoxalase 1. The glyoxalase-2-deficient red cells were used to test whether S-lactoylglutathione is transported from red cells via the glutathione-S-conjugate transporter; this transport appeared not to occur. Because methylglyoxal is extremely rapidly removed (half-life, approximately 5 min) from normal red cells, it is difficult to assess the effect of this compound on glycolysis but the slow decline evident in the deficient cells allowed a study of the effects on L-lactate production; no effects were apparent.
Rae
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Department of Biochemistry, University of Sydney, New South Wales, Australia.
Board
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PG
Kuchel
P W
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D000818:Animals; D001692:Biological Transport; D003198:Computer Simulation; D004912:Erythrocytes / Q000201:enzymology*; D005978:Glutathione / Q000031:analogs & derivatives* / Q000378:metabolism; D006736:Horses; D007700:Kinetics; D007791:Lactoylglutathione Lyase / Q000172:deficiency* / Q000378:metabolism; D007962:Leukocytes / Q000201:enzymology; D009682:Magnetic Resonance Spectroscopy; D011765:Pyruvaldehyde / Q000494:pharmacology
D011765:Pyruvaldehyde; D007791:Lactoylglutathione Lyase; D005978:Glutathione; C013585:S-lactoylglutathione