Nature and biosynthesis of sialic acids in the starfish Asterias rubens. Identification of sialo-oligomers and detection of S-adenosyl-L-methionine: N-acylneuraminate 8-O-methyltransferase and CMP-N-acetylneuraminate monooxygenase activities.

ABSTRACT

Mass spectrometric and NMR spectroscopic analyses of bound sialic acids from the starfish Asterias rubens revealed the presence of N-acetylneuraminic acid (4%), N-acetyl-8-O-methylneuraminic acid (12%), N-acetyl-9-O-acetyl-8-O-methylneuraminic acid (less than 1%), N-glycoloylneuraminic acid (19%), N-glycoloyl-8-O-methylneuraminic acid (47%), and N-glycoloyl-9-O-acetyl-8-O-methylneuraminic acid (18%). Analysis of sialo-oligomeric material, obtained after mild acid hydrolysis, demonstrated that N-glycoloyl-8-O-methylneuraminic acid can occur as di- and tri-oligomers, linked through the anomeric center and the N-glycoloyl moiety, Neu5Gc8Me-alpha(2----O5)-Neu5Gc8Me and Neu5Gc8Me-alpha(2----O5)-Neu5Gc8Me-alpha (2----O5)-Neu5Gc8Me. Studies on the biosynthesis of N-acyl-8-O-methylneuraminic acid in A rubens, using the tracer S-adenosyl-L-[methyl-14C]methionine, showed that N-acylneuraminate 8-O-methyltransferase activity was present predominantly in the membrane fraction. CMP-N-acetylneuraminic acid monooxygenase activity was found in the soluble protein fraction, in agreement with investigations on the corresponding vertebrate enzyme.