The Metabolism of O‐Acyl‐N‐acylneuraminic Acids

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Abstract
1 The enzymic synthesis of 4-O-acetyl-N-acetylneuraminic acid, 4-O-acetyl-N-glycolylneuraminic acid, 4-O-glycolyl-N-acetylneuraminic acid, 9-O-acetyl-N-acetylneuraminic acid and 9-O-acetyl-N-glycolylneuraminic acid is shown using radioactive precursors with surviving slices, membrane fractions or particle-free homogenates from bovine and equine submandibular glands. 2 Acetyl-CoA: N-acetylneuraminate-9(or 7)-O-acetyltransferase activity was found in a microsome fraction and in the cytosol of bovine submandibular glands. The properties of the membrane-bound enzyme acting on endogenous, glycoprotein-bound N-acetyl- and N-glycolylneuraminic acids were compared with those of the soluble enzyme, O-acetylating exogenous, non-glycosidically bound N-acetyl- and N-glycolylneuraminic acids. 3 A rapid, radioactive assay for the membrane-bound enzyme activity is described. The enzyme activity shows an optimum at pH 7 and has a Km for acetyl-CoA of 0.1 mM. The enzyme is inhibited by p-chloromercuribenzoate and iodoacetate. Divalent cations, EDTA and glutathione have no influence on its activity while CoA proved to be a competitive inhibitor with a K1 of 0.56 mM. 4 The soluble enzyme activity, assayed using a radioactive procedure, shows Km values of 0.01 mM, 0.5 mM and 0.39 mM for acetyl-CoA, N-acetylneuraminic acid and N-glycolylneuraminic acid respectively. The general properties are similar to those found for the membrane-bound enzyme, except that membrane-bound activity is stable for longer on storage at 4 °C. 5 Acetyl-CoA, acyl-CoA and CoA concentrations of 33 nmol, 65 nmol and 106 nmol/g wet tissue respectively are found in fresh bovine submandibular glands. 6 The occurrence of the CMP-glycosides of N-acetylneuraminic acid, 9-O-acetyl-N-acetylneuraminic acid and N-glycolylneuraminic acid in bovine submanibular glands is demonstrated. 7 The results are discussed in relation to the general metabolism of acylneuraminic acids.

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