Molecularly cloned mammalian glucosamine-6-phosphate deaminase localizes to transporting epithelium and lacks oscillin activity

^a Department of Neuroscience, The Johns Hopkins University, School of Medicine, Baltimore, Maryland 21205, USA
^b Department of Pharmacology and Molecular Sciences, The Johns Hopkins University, School of Medicine, Baltimore, Maryland 21205, USA
^c Department of Psychiatry and Behavioral Sciences, The Johns Hopkins University, School of Medicine, Baltimore, Maryland 21205, USA
^d Department of Biological Sciences, Kent State University, Kent, Ohio 41242, USA

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-phosphate. Though first identified in mammalian tissues in 1956, the enzyme has not previously been molecularly characterized in mammalian tissues, although a bacterial GNPDA has been cloned. Recently, a protein displaying similarity to bacterial GNPDA was purified and cloned from sperm extract. It was proposed that this protein was the factor, found in sperm extracts, that causes calcium oscillations in cells; thus, the protein was named 'oscillin.' We demonstrate that oscillin is the mammalian form of glucosamine 6-phosphate deaminase by showing that cloned oscillin has a robust GNPDA activity and can account for all such activity in mammalian tissues extracts. In situ hybridization and immunohistochemistry 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 homogeneity from hamster sperm fail to elevate intracellular calcium when injected into mouse eggs over a wide range of concentrations under conditions in which sperm extracts elicit pronounced calcium oscillations. Thus, the calcium-releasing or oscillin activity of sperm extracts is due to a substance other than GNPDA. Since GNPDA is the sole enzyme linking hexosamine systems with glycolytic pathways, we propose that it provides a source of energy in the form of phosphosugar derived from the catabolism of hexosamines found in glycoproteins, glycolipids, and sialic acid-containing macromolecules. Evidence that GNPDA can regulate hexosamine stores comes from our observation that transfection of GNPDA into HEK-293 cells reduces cellular levels of sialic acid. —Wolosker, H., Kline, D., Bian, Y., Blackshaw, S., Cameron, A. M., Fralich, T. J., Schnaar, R. L., Snyder, S. H. Molecularly cloned mammalian glucosamine-6-phosphate deaminase localizes to transporting epithelium and lacks oscillin activity. FASEB J. 12, 91–99 (1998)

Key Words: metabolism • hexosamines • cloning • sialic acid • GNPDA

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