Abstract
(LDH) obtained from larvae of Drosophila melanogaster was purified to homogeneity by affinity chromatography on oxamate-Sepharose. The purification procedure is simple to operate and gives a homogeneous preparation in a good yield (34.86%) after only two steps. Utilizing the homogeneous LDH preparation, an attempt was made to characterize the LDH molecule. Thus, it was found that the N-terminal amino acid is isoleucine, and the enzyme is tetrameric and composed of four identical subunits of apparent molecular weight 38,000, suggesting that it is controlled by a single gene. Homogeneous LDH preparations exhibit one band on neutral acrylamide gels when the substrate is either dl-lactic acid or l-(+)-lactate. The optimum temperature is 45°C for the purified enzyme and 40°C for the crude homogenate. The K m values for pyruvate and NADH are 0.154 and 0.027mm, respectively, while the K m values for lactate and NAD are 29.4 and 1.33mm, respectively. A discontinuity in the E a slope was observed at a transition temperature of 30°C. The E a value between 20 and 30°C was calculated as 12.06 kcal/mol, while between 30 and 45°C the E a value was 4.01 kcal/mol. This evidence, together with other observations reported in the literature, suggests that the LDHs of invertebrates and vertebrates have arisen by divergent evolution from a common ancestral gene.
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Onoufriou, A., Alahiotis, S.N. Drosophila lactate dehydrogenase: Molecular and genetic aspects. Biochem Genet 20, 1195–1209 (1982). https://doi.org/10.1007/BF00498943
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DOI: https://doi.org/10.1007/BF00498943