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Metabolism and Enzymology

Species and Environmental Variations in the Effect of Inorganic Phosphate on Sucrose-Phosphate Synthase Activity ¹

Reliability of Assays Based Upon UDP Formation

U.S. Department of Agriculture, Agricultural Research Service, University of Kentucky, Lexingon, Kentucky 40546

The effect of inorganic phosphate (Pi) on sucrose-phosphate synthase (SPS) activity was determined for the enzyme from five plant species (Nicotiana tabacum L., Spinacia oleracea L., Triticum aestivum L., Zea mays L., Glycine max L.) using two assay methods. The assay method based on determination of uridine diphosphate glucose- (UDPG) and fructose-6-phosphate-dependent sucrose formation was linear up to 15 minutes for all species tested. When assayed in this way, the effect of Pi at levels of 5 or 10 millimolar in the assay was variable, ranging from 0 to 35% inhibition of SPS activity. The assay method based on substrate dependent UDP formation was linear for some, but not for all of the species tested. Deviations from linearity were caused by loss of UDP from the assay medium. In some species, the extent of UDP loss was influenced by the level of Pi in the assay medium and, for at least one species (tobacco), it was influenced by the environment in which the plants were grown. The results indicated that (a) the role of Pi as an effector of SPS may vary depending on the species, and (b) the UDP assay method should be used with caution for assays of crude or desalted extracts, particularly when evaluating the effect of Pi on SPS activity.