Sialidases purification

H. Umezawa, T. Aoyagi, T. Komiyama, H. Morishima, M. Hamada, and T. Takeuchi, Purification and characterization of a sialidase inhibitor, siastatin, produced by Streptomyces, J. Antibiot., 27 (1974) 963-969. 

Only a few bacterial and viral sialidases have been purified to high purity or even to protein homogeneity."0 Complete purification of sia-lidase on a preparative scale from the culture filtrate ofC. perfringens was achieved111 by using poly(acrylamide) gel-electrophoresis as the final purification step (see Section VI,1). It is necessary that such purified sialidases be available, as the presence of proteases or other gly-cosidases in the enzyme preparations would lead to severe errors, not only in studies of substrate specificity, but also in cell biological and medical studies (see Sections VI and VII). 

Exact analysis of sialic acid is required in biologieal experiments where the biological role of sialic acid is frequently studied with the aid of sialidases, and the amount of sialic acids released is determined. This is also important for periodate oxidation studies on biological systems, where modification of sialic acids by periodate is only assumed, but chemical analysis of this effect by isolation and analysis of the modified sialic acids is seldom performed. These uncertainties in determinations of sialic acid can be overcome by the purification procedures already described. Furthermore, it must be stressed that unequivocal determination of the structure of a sialic acid, especially... 

Purification of a variety of viral sialidases (influenza and paramyxovirus) after solubilization by proteases or detergents has been exten-... 

Although most of the sialidases have been purified by classical methods of enzyme isolation, affinity chromatography is now coming more and more into use, as is indicated by the foregoing examples of purification of sialidases. The adsorbent first used for chromatography of sialidases was sialic acid bound to the surface of intact erythrocytes ... 

Chang, Y. K., and Chein, C. H. (1998). Simple two step procedure for purification of cloned small sialidase from unclaridied E. coli feedstocks, lnt. Conf. Expanded Bed Adsorption, 2nd, Napa Valley, CA, 1998, Abstr., p. 8.2. 

The peanut Arachis hypogaea) contains a lectin with anti-T (Gal(Pl-3)GalNAc) activity [174]. This antigen appears on human erythrocytes following treatment with sialidase and leads to the phenomenon known as polyagglutinability as monitored by the peanut lectin [175]. Peanut agglutinin, purified by numerous affinity purification schemes, is a tetrameric protein composed of four carbohydrate-free subunits, A/f = 27000Da[176]. The lectin is a metalloprotein rich in acidic and hydroxylic amino acids and devoid of cysteine [176]. 

M. Engstler, G. Reuter, and R. Schauer, Purification and characterization of a novel sialidase found in procycUc culture forms of Trypanosoma brucei. Mol Biochem. Parasitol, 54 (1992) 21-30. 

The purification and characterization of siastatin, a sialidase inhibitor produced by a Streptomyces species, have been described. ... 

The sialic acid preparations obtained with the aid of sialidase generally are purer than those obtained by acid hydrolysis. In spite of this, ultrafiltration or dialysis ( 10,000 daltons cut ofO is advisable before further purification. 

Viral sialidases are membrane components, and as such show greater thermal stability relative to bacterial enzymes. They require purification after release from the virus. Complexes in the range of 1-2 x 105 daltons, or monomers in the range of 5-6 X 104 daltons have been reported for sialidase preparations without proteolytic degradation (see Drzeniek 1972, Rosenberg and Schengrund 1976). 

J.T. Cassidy, G.W. Jourdian and S. Roseman, The sialic acid. VI. Purification and properties of sialidase from Clo.stridium perfringens. J. Biol Chem., 1965, 240, 3501-3506. 

L.L. Hoyer, P. Roggentin, R. Schauer and E.R. Vimr, Purification and properties of cloned Salmonella typhimurium LT2 sialidase with virus-typical kinetic preference for sialyl a 2,3 linkages. J. Biochem. (Tokyo), 1991, llO, 462-467. 

Heuermann, D., Roggentin, P., Kleineidam, R. G., and Schauer, R., 1991, Purification and characterization of a sialidase from Clostridium chauvoei NC08596, Glycoconj. J. 8 95-101. 

Tanaka, H., Ito, F., and lawasaki, T, 1992, Purification and characterization of a sialidase from Bacteroides fragilis SBT3182, Biochem. Biophys. Res. Commun. 189 524-529. 

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