An endogenous proteinacious inhibitor in porcine liver for S-adenosyl-L-methionine dependent methylation reactions: identification as oligosaccharide-linked acyl carrier protein

A proteinacious inhibitor of S-adenusyl-L-methionine (AdoMet)-dependent tra nsmethylation reactions was purified to homogeneity from porcine liver by s ize exclusion chromatography and FPLC. The molecular weight of the inhibito r was 12,222 Da, A 7400 Da polypeptide fragment of the purified inhibitor w as sequenced by matrix-associated laser desorption ionization: time-of-flig ht MS, and was found to be identical with the known sequence of spinach acy l carrier protein (ACP). Although the remainder of the molecule was not cle arly defined,H-1 and H-H correlation of spectroscopy; (COSY) NMR analysis r evealed the presence of an oligosaccharide with alpha-glycosidic linkage. T he purified oligosaccharide-linked ACP inhibited several AdoMet-dependent t ransmethylation reactions such as protein methylase I and II, S-farnesylcys teine O-methyltransferase, DNA methyltransferase and phospholipid methyltra nsferase. Protein methylase II was inhibited with a K-i value of 2.4 x 10(- 3) M in a mixed inhibition pattern. whereas a well-known competitive produc t inhibitor S-adenosyl-L-homocysteine (AdoHcy) had K-i value of 6.3 x 10(-6 ) M, Commercially available active ACP fragments (65-74) and ACP from Esche richia coli had less inhibitory activity toward S-farnesylcysteine O-methyl transferase than the purified inhibitor, The biological significance of thi s oligosaccharide-linked ACP which has two seemingly unrelated functions (i nhibitor for transmethylation and fatty acid biosynthesis) remains to be el ucidated, (C) 2000 Elsevier Science Ltd, All rights reserved.