STIMULATION OF TYROSYLPROTEIN SULFOTRANSFERASE ACTIVITY BY ETHANOL - ROLE OF INCREASED ENZYME LEVEL

Tyrosylprotein sulfotransferase (TPST), an enzyme involved in the post translational modification of proteins, plays important role in the bi ological activity and secretion of proteins. Previously we have shown an increased activity of this enzyme in gastric mucosa of alcoholics. In the present study, effect of ethanol on TPST was examined in rat li ver and gastric mucosa utilizing enzyme assays and Western blot analys es for TPST levels. Male Sprague-Dawley rats were pair-fed Leiber-DeCa rli liquid diets for 10 days and controls received a liquid diet in wh ich dextrose was isocalorically substituted for ethanol. After ethanol feeding, rats were sacrificed and liver and gastric mucosa were proce ssed for Golgi membrane preparation. The TPST activity was measured us ing poly(Glu(6), Ala(3), Tyr(1)) as the sulfate acceptor and PAPS as s ulfate donor. There was a threefold increase in TPST activity of gastr ic mucosa of animals subjected to chronic alcohol feeding. In the live r, the increase in tyrosine sulfating activity was also around threefo ld. The kinetic studies performed to understand the mechanism involved in ethanol stimulation of TPST activity showed no change in the K-m v alues of the enzyme by ethanol. In control and ethanol-treated animals , the K-m for EAY was 0.41-0.53 and 0.43-0.53 mu M, and the K-m for PA PS was 10-12.5 and 9-17 mu M, respectively. The V-max in ethanol-fed a nimals was increased by 1.5- to 2-fold. The increase in TPST activity in experimental rats was further tested by analyzing the Western blots by Imaging Densitometer for TPST levels. Analysis of TPST levels also showed over threefold increase in the stomach and liver of ethanol-fe d rats. Our results indicate that stimulation of TPST by ethanol invol ves increased TPST level rather than change in affinity for its substr ates. (C) 1998 Elsevier Science Inc.