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Isolation and properties of two sialate-O-acetylesterases from horse liver with 4- and 9-O-acetyl specificities

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Abstract

Sialate-O-acetylesterase was purified almost 900-fold from particle-free supernatants of horse liver by gel filtration, ion-exchange chromatography and isoelectric focussing. The native enzyme on gel filtration exhibits a molecular weight of 54,000 Da. It was separated by isoelectric focussing into two forms with pI values of 4.8 and 5.7, respectively. The esterase with a lower pI hydrolyses only 9-O-acetyl groups from sialic acids (KM 1.1 mM), while that with the higher pI esterifies both 4- and 9-O-acetylated monosaccharides at similar rates (KM 0.3 M and 1.3 mM, respectively). Both forms are inactive with 7-O-acetylated N-acetylneuraminic acid. Enzyme assays were carried out at the pH optimum (pH 8.4–8.6) using free O-acetylated sialic acids followed by direct analysis of the reaction products by isocratic anion-exchange HPLC. Glycosidically bound sialic acids can also be de-O-acetylated. Horse liver esterase seems to be an essential enzyme for the catabolism of 4-O-acetylated sialoglycoconjugates, since sialidase from this tissue cannot act on 4-O-acetylated sialic acids.

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Acknowledgements

The authors thank Sabine Stoll and Margret Wember for excellent technical assistance, and G. Vinayaga Srinivasan for help in preparing the manuscript. The financial support by Deutsche Forschungsgemeinschaft (grant Scha 202/10-4) and the Fonds der Chemischen Industrie is greatfully acknowledged. Thanks are also due to Prof. Dr. E. Heymann for discussion.

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  1. Biochemisches Institut, Christian-Albrechts-Universität, Olshausenstr. 40, 24098, Kiel, Germany

    Roland Schauer

  2. Glygen Corp., Columbia, MD, 21045, USA

    Ashok K. Shukla

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Schauer, R., Shukla, A.K. Isolation and properties of two sialate-O-acetylesterases from horse liver with 4- and 9-O-acetyl specificities. Glycoconj J 25, 625–632 (2008). https://doi.org/10.1007/s10719-008-9109-9

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