A Unique Sialidase That Cleaves the Neu5Gcα2→5-OglycolylNeu5Gc Linkage: Comparison of Its Specificity with That of Three Microbial Sialidases toward Four Sialic Acid Dimers
Biochemical and Biophysical Research Communications(2001)
摘要
We found that the hepatopancreas of oyster, Crassostrea virginica, contained a sialidase capable of releasing Neu5Gc from the novel polysialic acid chain (→5-OglycolylNeu5Gcα2→)n more efficiently than from the conventional type of polysialic acid chains, (→8Neu5Acα2→)n, or (→8Neu5Gcα2→)n. We have partially purified this novel sialidase and compared its reactivity with that of microbial sialidases using four different sialic acid dimers, Neu5Gcα2→5-OglycolylNeu5Gc (Gg2), Neu5Acα2→8Neu5Ac (A2), Neu5Gcα2→8Neu5Gc (G2), and KDNα2→8KDN (K2) as substrates. Hydrolysis was monitored by high performance anion-exchange chromatography with a CarboPac PA-100 column and pulsed amperometric detection, the method by which we can accurately quantitate both the substrate (sialiac acid dimers) and the product (sialic acid monomers). The oyster sialidase effectively hydrolyzed Gg2 and K2, whereas A2 and G2 were poor substrates. Neu5Ac2en but not KDN2en effectively inhibited the hydrolysis of Gg2 by the oyster sialidase. Likewise, the hydrolysis of K2 by the oyster sialidase was inhibited by a cognate inhibitor, KDN2en, but not by Neu5Ac2en. Using the new analytical method we found that Gg2 was hydrolyzed less efficiently than A2 but much more readily than G2 by Arthrobacter ureafaciens sialidase. This result was at variance with the previous report using the thiobarbituric acid method to detect the released free sialic acid [Kitazume, S., et al. (1994) Biochem. Biophys. Res. Commun. 205, 893–898]. In agreement with previous results, Gg2 was a poor substrate for Clostridium perfringens sialidase, while K2 was refractory to all microbial sialidases tested. Thus, the oyster sialidase is novel and distinct from microbial sialidases with regards to glycon- and linkage-specificity. This finding adds an example of the presence of diverse sialidases, in line with the diverse sialic acids and sialic acid linkages that exist in nature. The new sialidase should become useful for both structural and functional studies of sialoglycoconjugates.
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关键词
sialidases,Neu5Gcα2→5-OglycolylNeu5Gc-specific sialidase,Arthrobacter ureafaciens sialidase,oyster sialidase
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