Optimization of efficiency in the glyoxalase pathway.
A quantitative kinetic model for the glutathione-dependent conversion of methylglyoxal to D-lactate in mammalian erythrocytes has been formulated, on the basis of the measured or calculated rate and equilibrium constants associated with (a) the hydration of methylglyoxal, (b) the specific base catalyzed formation of glutathione-(R,S)-methylglyoxal thiohemiacetals, (c) the glyoxalase I catalyzed conversion of the diastereotopic thiohemiacetals to (S)-D-lactoylglutathione, and (d) the glyoxalase II catalyzed hydrolysis of (S)-D-lactoylglutathione to form D-lactate and glutathione. The model exhibits the following properties under conditions where substrate concentrations are small in comparison to the Km values for the glyoxalase enzymes: The overall rate of conversion of methylglyoxal to D-lactate is primarily limited by the rate of formation of the diastereotopic thiohemiacetals. The hydration of methylglyoxal is kinetically unimportant, since the apparent rate constant for hydration is (approximately 500-10(3))-fold smaller than that for formation of the thiohemiacetals. The rate of conversion of methylglyoxal to (S)-D-lactoylglutathione is near optimal, on the basis that the apparent rate constant for the glyoxalase I reaction (kcatEt/Km congruent to 4-20 s-1 for pig, rat, and human erythrocytes) is roughly equal to the apparent rate constant for decomposition of the thiohemiacetals to form glutathione and methylglyoxal [k(obsd) = 11 s-1, pH 7]. The capacity of glyoxalase I to use both diastereotopic thiohemiacetals, versus only one of the diastereomers, as substrates represents a 3- to 6-fold advantage in the steady-state rate of conversion of the diastereomers to (S)-D-lactoylglutathione.(ABSTRACT TRUNCATED AT 250 WORDS)
Creighton
D J
DJ
Department of Chemistry, University of Maryland Baltimore County, Catonsville 21228.
Migliorini
M
M
Pourmotabbed
T
T
Guha
M K
MK
D000818:Animals; D004912:Erythrocytes / Q000201:enzymology*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes / Q000097:blood; D007700:Kinetics; D007773:Lactates / Q000097:blood; D019344:Lactic Acid; D007791:Lactoylglutathione Lyase / Q000097:blood*; D008190:Lyases / Q000097:blood*; D008623:Mercaptoethanol / Q000378:metabolism; D011765:Pyruvaldehyde / Q000097:blood; D051381:Rats; D013237:Stereoisomerism; D013379:Substrate Specificity; D013552:Swine; D013816:Thermodynamics; D013864:Thioglycolates / Q000378:metabolism; D013869:Thiolester Hydrolases / Q000097:blood*; D000099301:Hydroxyacylglutathione Hydrolase
D007527:Isoenzymes; D007773:Lactates; D019344:Lactic Acid; D007791:Lactoylglutathione Lyase; D008190:Lyases; D008623:Mercaptoethanol; D011765:Pyruvaldehyde; D013864:Thioglycolates; D013869:Thiolester Hydrolases; D000099301:Hydroxyacylglutathione Hydrolase; C037474:methyl thioglycolate