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J. Biol. Chem., Vol. 278, Issue 26, 23301-23310, June 27, 2003

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Two Trans-sialidase Forms with Different Sialic Acid Transfer and Sialidase Activities from Trypanosoma congolense^*

Evelin Tiralongo, Silke Schrader , Hans Lange, Hilmar Lemke, Joe Tiralongo  and Roland Schauer ¶

From the Biochemisches Institut, Universität zu Kiel, Olshausenstrasse 40, Kiel 24098, Germany

Trypanosomes express an enzyme called trans-sialidase (TS), which enables the parasites to transfer sialic acids from the environment onto trypanosomal surface molecules. Here we describe the purification and characterization of two TS forms from the African trypanosome Trypanosoma congolense. The purification of the two TS forms using a combination of anion exchange chromatography, isoelectric focusing, gel filtration, and subsequently, antibody affinity chromatography resulted, in both cases, in the isolation of a 90-kDa monomer on SDS-PAGE, which was identified as trans-sialidase using micro-sequencing. Monoclonal antibody 7/23, which bound and partially inhibited TS activity, was found in both cases to bind to a 90-kDa protein. Both TS forms possessed sialidase and transfer activity, but markedly differed in their activity ratios. The TS form with a high transfer-to-sialidase activity ratio, referred to as TS-form 1, possessed a pI of pH 4–5 and a molecular mass of 350–600 kDa. In contrast, the form with a low transfer-to-sialidase activity ratio, referred to as TS-form 2, exhibited a pI of pH 5–6.5 and a molecular mass of 130–180 kDa. Both TS forms were not significantly inhibited by known sialidase inhibitors and revealed no significant differences in donor and acceptor substrate specificities; however, TS-form 1 utilized various acceptor substrates with a higher catalytic efficiency. Interestingly, glutamic acid-alanine-rich protein, the surface glycoprotein, was co-purified with TS-form 1 suggesting an association between both proteins.

Received for publication, December 18, 2002 , and in revised form, March 19, 2003.

^* This work was financially supported by the German Federal Ministry of Education and Research (Project 0311827A) and Numico Research, Germany, as well as by the Fonds der Chemischen Industrie (Frankfurt) and the Sialic Acids Society (Kiel). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. AJ535487 and AJ535488.

Current address: Institut für Biochemie, Universität zu Köln, Zülpicher Strasse 47, Köln 50674, Germany.

Current address: Zentrum Biochemie, Abteilung Zelluläre Chemie, Medizinische Hochschule Hannover, Carl-Neuberg-Strasse 1, Hannover 30625, Germany.

^¶ To whom correspondence should be addressed. Tel.: 49-431-880-2210; Fax: 49-431-880-2238; E-mail: schauer{at}biochem.uni-kiel.de.

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