The occurrence of 4-methylthio-2-hydroxybutyrate in human urine.
A method for determination of 4-methylthio-2-hydroxybutyrate and 4-methylthio-2-oxobutyrate in human urine has been devised, based on metoxime formation of the keto acid and a clean-up procedure using a strong anion-exchange resin AG 2 X 8 and ethyl acetate extraction. After alkylation, the compounds were quantified by GC, using a flame photometric sulfur-selective detector. A normal urinary excretion of 0.14 to 0.25 mmol/mol creatinine and 0.07 to 0.22 mmol/mol creatinine of the alpha-hydroxy and alpha-keto acid, respectively, was found, whereas a markedly elevated excretion of the hydroxy acid was noted in subjects with hypermethioninemia. The enzymatic reduction of 4-methylthio-2-oxobutyric acid by lactate dehydrogenase: NAD+ oxidoreductase (EC 1.1.1.17) was also studied. The Km and Kequil values for 4-methylthio-2-oxobutyrate were 1.41 mM and 0.92 X 10(8) M-1. The Vmax value of the enzyme at infinite concentrations of the two substrates was 7.2 mumol/s/mumol enzyme, which indicates low affinity and reduced catalytic activity compared to other known substrates of lactate dehydrogenase. The reaction product 4-methylthio-2-hydroxybutyrate was not inhibitory on the reaction. The M4 isoenzyme of lactate dehydrogenase (rabbit and pig muscle) possessed approximately 20% of the activity of the H4 isoenzyme (pig heart) for the substrate.
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J
Mårtensson
D000818:Animals; D006801:Humans; D006885:Hydroxybutyrates; D066298:In Vitro Techniques; D007527:Isoenzymes / Q000378:metabolism; D007700:Kinetics; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008715:Methionine / Q000031:analogs & derivatives* / Q000378:metabolism / Q000652:urine; D009132:Muscles / Q000201:enzymology; D009206:Myocardium / Q000201:enzymology; D011817:Rabbits; D013379:Substrate Specificity; D013552:Swine
D006885:Hydroxybutyrates; D007527:Isoenzymes; C008391:alpha-hydroxy-gamma-methylmercaptobutyric acid; C014632:2-keto-4-methylthiobutyric acid; D008715:Methionine; D007770:L-Lactate Dehydrogenase