Comparison of in vitro and in vivo activities associated with the G6PD allozyme polymorphism in Drosophila melanogaster.
Earlier studies of the A and B allozymes at the G6pd locus show a differential ability of the genotypes to suppress the loss of viability associated with a low activity 6-phosphogluconate dehydrogenase mutation, 6Pgdlo1. This observation indicates a relatively lower activity for the A allozyme genotype, but it is not known if this level of suppression required a large difference in in vivo activity. To clarify this difference an analysis of the biochemical properties of the purified allozymes was carried out, as well as an analysis of the activity level associated with an original low activity P element-derived allele which had partially reverted and lost its suppression ability. G6PD activity and protein level were studied in 47 X chromosome lines from North America. The A genotype averages a 9% lower Vmax. From analysis of the correlation between G6PD activity and protein level it remains unclear whether the allozyme Vmax difference results from dissimilarity in protein level or kcat. At 25 degrees and physiological pH, comparative studies of the steady-state kinetics show the two purified allozyme variants differ significantly in their KM values for glucose-6-phosphate and NADP, and the K1 for NADPH. In aggregate these parameters predict the A genotype possesses a 20% lower in vitro catalytic efficiency. A partial revertant of a P element-derived low activity B variant, was shown to lose the ability to suppress 6Pgdlo1 low viability after acquiring only 60% of normal B activity. This last comparison shows the A genotype activity must be reduced in vivo by at least 40%.
Eanes
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WF
Department of Ecology and Evolution, State University of New York, Stony Brook 11794.
Katona
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L
Longtine
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M
D000704:Analysis of Variance; D000818:Animals; D003433:Crosses, Genetic; D004251:DNA Transposable Elements; D004331:Drosophila melanogaster / Q000201:enzymology / Q000235:genetics*; D005838:Genotype; D005954:Glucosephosphate Dehydrogenase / Q000235:genetics* / Q000378:metabolism; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D009249:NADP / Q000378:metabolism; D010734:Phosphogluconate Dehydrogenase / Q000235:genetics / Q000378:metabolism; D011110:Polymorphism, Genetic; D015183:Restriction Mapping; D013489:Suppression, Genetic; D013696:Temperature
D004251:DNA Transposable Elements; D007527:Isoenzymes; D009249:NADP; D010734:Phosphogluconate Dehydrogenase; D005954:Glucosephosphate Dehydrogenase