The Aspergillus fumigatus Sialidase Is a 3-Deoxy-D-glycero-D-galacto-2-nonulosonic Acid Hydrolase (KDNase): Structural and Mechanistic Insights
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Aspergillus fumigatus is a filamentous fungus that can cause severe respiratory disease in immunocompromised individuals. A putative sialidase from A. fumigatus was recently cloned and shown to be relatively poor in cleaving N-acetylneuraminic acid (Neu5Ac) in comparison with bacterial sialidases. Here we present the first crystal structure of a fungal sialidase. When the apo structure was compared with bacterial sialidase structures, the active site of the Aspergillus enzyme suggested that Neu5Ac would be a poor substrate because of a smaller pocket that normally accommodates the acetamido group of Neu5Ac in sialidases. A sialic acid with a hydroxyl in place of an acetamido group is 2-keto-3-deoxynononic acid (KDN). We show that KDN is the preferred substrate for the A. fumigatus sialidase and that A. fumigatus can utilize KDN as a sole carbon source. A 1.45-Šresolution crystal structure of the enzyme in complex with KDN reveals KDN in the active site in a boat conformation and nearby a second binding site occupied by KDN in a chair conformation, suggesting that polyKDN may be a natural substrate. The enzyme is not inhibited by the sialidase transition state analog 2-deoxy-2,3-dehydro-N-acetylneuraminic acid (Neu5Ac2en) but is inhibited by the related 2,3-didehydro-2,3-dideoxy-D-glycero-D-galacto-nonulosonic acid that we show bound to the enzyme in a 1.84-Šresolution crystal structure. Using a fluorinated KDN substrate, we present a 1.5-Šresolution structure of a covalently bound catalytic intermediate. The A. fumigatus sialidase is therefore a KDNase with a similar catalytic mechanism to Neu5Ac exosialidases, and this study represents the first structure of a KDNase.
Journal of Biological Chemistry
This research was originally published in Journal of Biological Chemistry (JBC). Judith C. Telford, Juliana H.F. Yeung, Guogang Xu, Milton J. Kiefel, Andrew G. Watts, Stefan Hader, Jefferson Chan, Andrew J. Bennet, Margo M. Moore and Garry L. Taylor,The Aspergillus fumigatus Sialidase Is a 3-Deoxy-D-glycero-D-galacto-2-nonulosonic Acid Hydrolase (KDNase): Structural and Mechanistic Insights, Journal of Biological Chemistry (JBC), 2011; Vol.286: pp.10783-10792. Copyright the American Society for Biochemistry and Molecular Biology. This is the author-manuscript version of this paper. Reproduced in accordance with the copyright policy of the publisher. Please refer to the journal's website for access to the definitve version.
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