MOLECULARLY CLONED MAMMALIAN GLUCOSAMINE-6-PHOSPHATE DEAMINASE LOCALIZES TO TRANSPORTING EPITHELIUM AND LACKS OSCILLIN ACTIVITY

Glucosamine-6-phosphate deaminase (GNPDA) catalyzes the conversion of glucosamine-6-phosphate to fructose-6-phosphate, a reaction that under physiological conditions proceeds to the formation of fructose-6-phos phate. Though first identified in mammalian tissues in 1956, the enzym e has not previously been molecularly characterized in mammalian tissu es, although a bacterial GNPDA has been cloned. Recently, a protein di splaying similarity to bacterial GNPDA was purified and cloned from sp erm extract. It was proposed that this protein was the factor, found i n sperm extracts, that causes calcium oscillations in cells; thus, the protein was named 'oscillin.' We demonstrate that oscillin is the mam malian form of glucosamine 6-phosphate deaminase by showing that clone d oscillin has a robust GNPDA activity and can account for all such ac tivity in mammalian tissues extracts. In situ hybridization and immuno histochemistry localize GNPDA selectively to tissues with high energy requirements such as the apical zone of transporting epithelia in the proximal convoluted tubules of the kidney and the small intestine; to neurons (but not glia) and especially to nerve terminals in the brain; and to motile sperm. Recombinant GNPDA and GNPDA purified to homogene ity from hamster sperm fail to elevate intracellular calcium when inje cted into mouse eggs over a wide range of concentrations under conditi ons in which sperm extracts elicit pronounced calcium oscillations. Th us, the calcium-releasing or oscillin activity of sperm extracts is du e to a substance other than GNPDA. Since GNPDA is the sole enzyme link ing hexosamine systems with glycolytic pathways, we propose that it pr ovides a source of energy in the form of phosphosugar derived from the catabolism of hexosamines found in glycoproteins, glycolipids, and si alic acid-containing macromolecules. Evidence that GNPDA can regulate hexosamine stores comes from our observation that transfection of GNPD A into HEK-293 cells reduces cellular levels of sialic acid.