Human LDH Research Articles

Biochemical properties of trypanosomatid lactate dehydrogenases.

Lactate dehydrogenase (LDH, E.C.1.1.1.27) was found in supernatant (cytoplasmic enzyme) fractions of the trypanosomatid flagellates Trypanosoma conorhini and Crithidia fasciculata if 10 mm cysteine was present in the homogenizing medium. The T. conorhini LDH activity with pyruvate as substrate was increased 35% if 5 mm cysteine was also included in reaction mixtures. K(m) values for the T. conorhini enzyme were 3.3 x 10(-4)m with pyruvate, and 1.6 x 10(-4)m with alpha-ketobutyrate. Cysteine inhibited alpha-ketobutyrate reduction. Comparison of trypanosomatid and human serum LDH enzymes with respect to K(m), substrate activity and inhibition, pH optima, and K(i) values for oxalate and oxamate indicated that the trypanosomatid isoenzymes differed significantly from serum LDH. C. fasciculata LDH was extremely labile, since 59% of the activity was lost 90 min after isolation. The role of LDH enzymes in trypanosomatid metabolism is discussed, and the results are related to other trypanosomatid LDH enzymes. The comparison of homologous enzymes in host and parasite is discussed with regard to metabolic function and a possible model system for chemotherapy.

A Comparison of Reduced-NAD Preparations from Four Commercial Sources

Abstract The physical and chemical properties of several commercial sources of reduced NAD were examined. Properties compared included physical appearance, sodium and phosphorus content, ultraviolet light absorption at 260 and 340 nm, and relative rates of oxidation with two human LDH isoenzymes. Major differences between individual preparations were noted with respect to physical and chemical properties, as well as in the activity measurements. The commercial reduced-NAD preparations most rapidly oxidized by LDH were white, free-flowing substances with 260 nm:340 nm absorbance (A260:A340) ratios below 2.45. Detectable amounts of an LDH inhibitor were found in all preparations examined. A strong LDH inhibitor, which had the same properties as the inhibitor present in commercial reduced NAD, was isolated by column chromatography. This LDH inhibitor was found to produce the same degree of inhibition toward four LDH isoenzymes.

Optimum Reaction Conditions for Human Lactate Dehydrogenase Isoenzymes as They Affect Total Lactate Dehydrogenase Activity

Abstract Optimum reaction conditions at 30° ± 0.5° for two continuous spectrophotometric assay procedures, lactate to pyruvate (L → P) and pyruvate to lactate (P → L), were determined with respect to pH at 30° (pH30) substrate concentration, and coenzyme concentration for the human LDH isoenzymes. For the P → L procedure, broad pH30 optima were within the range of 7.20-7.40 for all the LDH isoenzymes. The coenzyme optima were identical for all of the isoenzymes tested at a reduced NAD concentration of 1.5 x 10-4 M. Pyruvate substrate optima ranged from 7.5 x 10-4 M for LDH1 to 1.7 x 10-3 M for LDH5 at pH30 7.30. For the L → P procedure, the pH30 optima were within the range of 8.30-8.88 for the LDH5 through LDH1 isoenzymes, respectively. Optimum activity was obtained at a NAD concentration of 6.0 x 10-3 M and remained constant at least to 1.8 x 10-2 M for each of the isoenzymes tested. L-Lactate substrate optima ranged from 4.0 x 10-2 M for LDH1 to at least 7.2 x 10-2 M for LDH5. From the isoenzyme studies, the degree of variation possibly involved in either method due to variations in isoenzyme distribution was calculated for total LDH samples. These calculations showed that both methods, P → L and L → P, were essentially equivalent. This equivalency was verified by a comparative study of the two methods on human serum samples.

Lactate dehydrogenase activity in high pyruvate concentrations modified by urea

The effect of urea on human LDH 1 and LDH 5 isoenzymes and on the LDH activities of human heart and liver homogenates was studied at various pyruvate concentrations. It was demonstrated that in high pyruvate concentrations (from 1.5 · 10 −3 M upward) the activity of LDH 1 and heart homogenate was greater in urea than when measured without urea. On the contrary, the activity of LDH 5 and liver homogenate was almost completely destroyed in urea. using a high pyruvate concentration (4 · 10 −3 M) the rather preliminary clinical results show that the serum LDH activity in patients with myocardial infarction is greater in urea than without urea. In liver diseases the serum LDH activity was lower in urea than without urea.

The effect of pyruvate on urea-induced inhibition of lactic dehydrogenase isozymes

By raising strongly the concentration of pyruvate we succeeded in elevating the activity of human LDH 1 isozyme up to 125% in the presence of urea compared with the standard made without urea. In the same circumstances human LDH 5 was inactivated to 6%. The activation-inactivation balance was studied with animal isozymes in different conditions. Clinical applicability of the test presented is outlined.