PURIFICATION AND SUBSTRATE KINETICS OF PLANT LACTATE-DEHYDROGENASE

Lactate dehydrogenase (LDH) from turnip (Brassica mpa; Cruciferae), pu rified to electrophoretic homogeneity using affinity chromatography, h as a native M-r of 157 x 10(3) and a subunit M-r of 38 x 10(3). The LD H from turnip shows the same relative effectiveness (relative V-max an d K-m values) as the mammalian H-4 and M-4 isoenzymes with pyruvate, l actate and glyoxylate (oxoacetate and dihydroxyacetate) as substrates. All three LDH types show no activity with glycolate (hydroxyacetate). The affinities for these and a range of competitive inhibitory analog ues shows a consistent pattern of highest affinity for the H-4 mammali an isoenzyme, medium affinity for the M-4 form and lowest affinity for the plant enzyme, in a ratio of about 10:3:1, respectively. The catal ytic mechanism of the plant enzyme is very similar to that of the mamm alian forms. The major physiological activity of the plant LDH is cons idered to be pyruvate reduction, rather than the disproportionation of glyoxylate that has been proposed as a plant cell pH-stat. (C) 1997 E lsevier Science Ltd. All rights reserved.