UDP-N-ACETYLGLUCOSAMINE TRANSFERASE AND GLUTAMINE - FRUCTOSE 6-PHOSPHATE AMIDOTRANSFERASE ACTIVITIES IN INSULIN-SENSITIVE TISSUES

Glutamine:fructose 6-phosphate amido-transferase (GFA) is rate-limitin g for hexosamine biosynthesis, while a UDP-GlcNAc beta-N-acetylglucosa minyltransferase (O-GlcNAc transferase) catalyses final O-linked attac hment of GlcNAc to serine and threonine residues on intracellular prot eins. Increased activity of the hexosamine pathway is a putative media tor of glucose-induced insulin resistance but the mechanisms are uncle ar. We determined whether O-GlcNAc transferase is found in insulin-sen sitive tissues and compared its activity to that of GFA in rat tissues . We also determined whether non-insulin-dependent diabetes mellitus ( NIDDM) or acute hyperinsulinaemia alters O-GlcNAc transferase activity in human skeletal muscle, O-GlcNAc transferase was measured using H-3 -UDP-GlcNAc and a synthetic cationic peptide substrate containing seri ne and threonine residues, and GFA was determined by measuring a fluor escent derivative of GlcN6P by HPLC. O-GlcNAc transferase activities w ere 2-4 fold higher in skeletal muscles and the heart than in the live r, which had the lowest activity, while GFA activity was 14-36-fold hi gher in submandibular gland and 5-18 fold higher in the liver than in skeletal muscles or the heart. In patients with NIDDM (n=11), basal O- GlcNAc transferase in skeletal muscle averaged 3.8 +/- 0.3 nmol/mg . m in, which was not different from that in normal subjects (3.3 +/- 0.4 nmol/mg . min). A 180-min intravenous insulin infusion (40 mU/m(2) . m in) did not change muscle O-GlcNAc transferase activity in either grou p. We conclude that O-GlcNAc transferase is widely distributed in insu lin-sensitive tissues in the rat and is also found in human skeletal m uscle. These findings suggest the possibility that O-linked glycosylat ion of intracellular proteins is involved in mediating glucose toxicit y. O-GlcNAc transferase does not, however, appear to be regulated by e ither NIDDM or acute hyperinsulinaemia, suggesting that mass action ef fects determine the extent of O-linked glycosylation under hyperglycae mic conditions.