Mechanisms for plasma-mediated activation of human blood cell aldehyde dehydrogenase.
Aldehyde dehydrogenase (ALDH; EC 1.2.1.3) activity assays were carried out on isolated human blood cells in phosphate-buffered saline (PBS) and in PBS mixed with human plasma. In assays with intact erythrocytes or sonicated leukocytes, the presence of 50% (v/v) or greater of plasma in the reaction mixtures produced a 2-fold increase in the rate of aldehyde oxidation. In corresponding assays with sonicated erythrocyte samples, the ALDH activity was enhanced on an average 1.5-fold, whereas a slight decrease was observed in assays with intact leukocytes. The ALDH inhibitor disulfiram almost completely abolished the enzyme activity both in the absence and presence of plasma. In assays with sonicated leukocytes, the activation effect could be antagonized by EDTA, indicating that it was caused largely by divalent cations. With sonicated erythrocytes, a significantly reduced ALDH activity was found only with the highest concentration of EDTA tested, and since a similar reduction was obtained also when plasma was omitted, the plasma-mediated activation of erythrocyte ALDH was suggested to be due to a different mechanism. After separation of plasma by gel filtration, an active fraction was identified by GC-MS and 1H-NMR to contain pyruvic acid, lactic acid and glucose. When tested at physiological plasma concentrations, pyruvic acid caused an increase in erythrocyte ALDH activity similar to that obtained with plasma, while lactic acid and glucose did not. Pyruvic acid did not activate the leukocyte ALDH. Based on these results, it is indicated that the plasma-mediated activation of erythrocyte ALDH is due to pyruvic acid, which reoxidizes NADH via lactate dehydrogenase (EC 1.1.1.27) and, thereby, increases the rate of dissociation of NADH from the terminal enzyme-NADH complex, the rate-limiting step in the ALDH pathway.
Helander
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Department of Zoophysiology, Uppsala University, Sweden.
Gogoll
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D000444:Aldehyde Dehydrogenase / Q000037:antagonists & inhibitors / Q000097:blood* / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D004912:Erythrocytes / Q000166:cytology / Q000201:enzymology*; D008401:Gas Chromatography-Mass Spectrometry; D005947:Glucose / Q000494:pharmacology; D006801:Humans; D007773:Lactates / Q000494:pharmacology; D019344:Lactic Acid; D007962:Leukocytes / Q000166:cytology / Q000201:enzymology*; D009682:Magnetic Resonance Spectroscopy; D010949:Plasma / Q000737:chemistry; D011773:Pyruvates / Q000494:pharmacology; D019289:Pyruvic Acid
D007773:Lactates; D011773:Pyruvates; D019344:Lactic Acid; D019289:Pyruvic Acid; D000444:Aldehyde Dehydrogenase; D005947:Glucose