Enzymatic studies of the prenatal chick, with special reference to the multibple forms of liver soluble NADP-linked isocitric dehydrogenase Public Deposited

http://ir.library.oregonstate.edu/concern/graduate_thesis_or_dissertations/8c97kv11j

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  • Levels of activity of glucose-6-phosphate- , 6-phosphogluconate- , and isocitric- dehydrogenase were measured in the soluble and particulate fractions of heart, brain and liver at several ages throughout the development of the chick embryo. Compared to isocitric dehydrogenase, levels of glucosesphosphate- , and 6-phosphogluconate- dehydrogenase are generally low. This suggests that the majority of the NADPH in the prenatal chick arises from the NADP-isocitric dehydrogenase. The results, however, say nothing of the possibility of different pools of NADPH arising from the different enzymes. A NADPH-NAD transhydrogenase activity was found in the digitonin-treated mitochondrial fractions of the same organs, and its levels were also determined. They were found to be very low, making it unlikely that the majority of the NADPH generated by the isocitric dehydrogenase is shunted into the electron transport chain of the mitochondria. Multiple forms of the NADP-linked isocitric dehydrogenase were shown to be present in the soluble fractions from liver of 18- day old embryos, using differential rates of heat denaturation. Partial purification of the soluble liver enzyme yielded two forms, one stable and the other labile to heat. It was found that these two enzymes have different sedimentation properties, pH optima and reaction rates with NADP. The dimeric, heat stable form may be dissociated into the monomeric, heat labile, form by a lowering of the ionic strength. This dissociation is readily reversible, and the newly formed dimer appears to be similar with the native dimer. On the basis of experiments presented and comparison with the behavior of similar enzymes in other organisms, a mode of interaction of the multiple forms is postulated. Soluble NADP-linked isocitric dehydrogenase of liver may play a role in the regulation of cellular metabolism during development.
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