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-adenosyl-L-methionine (AdoMet)-dependent transmethylation reactions was purified to homogeneity from porcine liver by size exclusion chromatography and FPLC. The molecular weight of the inhibitor was 12,222 Da. A 7400 Da polypeptide fragment of the purified inhibitor was sequenced by matrix-associated laser desorption ionization; time-of-flight MS, and was found to be identical with the known sequence of spinach acyl carrier protein (ACP). Although the remainder of the molecule was not clearly defined, 1H and H-H correlation of spectroscopy (COSY) NMR analysis revealed the presence of an oligosaccharide with alpha-glycosidic linkage. The purified oligosaccharide-linked ACP inhibited several AdoMet-dependent transmethylation reactions such as protein methylase I and II. S-farnesylcysteine O-methyltransferase, DNA methyltransferase and phospholipid methyltransferase. Protein methylase II was inhibited with a Ki value of 2.4 x 10(-3) M in a mixed inhibition pattern, whereas a well-known competitive product inhibitor S-adenosyl-L-homocysteine (AdoHcy) had Ki value of 6.3 x 10(-6) M. Commercially available active ACP fragments (65-74) and ACP from Escherichia coli had less inhibitory activity toward S-farnesylcysteine O-methyltransferase than the purified inhibitor. The biological significance of this oligosaccharide-linked ACP which has two seemingly unrelated functions (inhibitor for transmethylation and fatty acid biosynthesis) remains to be elucidated.
Seo
D W
DW
Department of Genetic Engineering, College of Life Science and Natural Resources, Sungkyunkwan University, Suwon, South Korea.
Moon
H I
HI
Han
J W
JW
Hong
S Y
SY
Lee
H Y
HY
Kim
S
S
Paik
W K
WK
Lee
H W
HW
D000213:Acyl Carrier Protein / Q000737:chemistry* / Q000302:isolation & purification; D000818:Animals; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000302:isolation & purification; D007700:Kinetics; D008099:Liver / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D008745:Methylation; D008780:Methyltransferases / Q000037:antagonists & inhibitors / Q000737:chemistry; D008970:Molecular Weight; D009844:Oligosaccharides / Q000737:chemistry*; D010446:Peptide Fragments / Q000737:chemistry; D011496:Protein Methyltransferases / Q000737:chemistry; D051381:Rats; D012436:S-Adenosylmethionine / Q000737:chemistry*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013552:Swine; D017855:Transferases (Other Substituted Phosphate Groups) / Q000737:chemistry
D000213:Acyl Carrier Protein; D004791:Enzyme Inhibitors; D009844:Oligosaccharides; D010446:Peptide Fragments; D012436:S-Adenosylmethionine; D008780:Methyltransferases; D011496:Protein Methyltransferases; C056501:protein-S-isoprenylcysteine O-methyltransferase; D017855:Transferases (Other Substituted Phosphate Groups); C021364:holo-(acyl-carrier-protein) synthase