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Purification and properties of a membrane-bound L-asparaginase of Tetrahymena pyriformis

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Abstract

L-Asparaginase activity reaches maximal values at the stationary phase of growth of Tetrahymena pyriformis and fluctuates upon the growth conditions and the composition of the medium. Most of the L-asparaginase activity (80%) is associated with the endoplasmic reticulum, and the remaining with the pellicles. Detergents either alone or in combination with NaCl up to 0.5 M concentration failed to solubilize L-asparaginase. Solubilization can be accomplished by means of either the chaotropic agents KSCN and NaClO4, or 0.1 M sodium phosphate buffer pH 8.0, following pretreatment of the particulates with 2% w/v Triton X100. L-Asparaginase has been purified to near homogeneity by hydrophobic and gel filtration chromatography. The native enzyme has a relative molecular weight of 230000. It is a multiple subunit enzyme, with subunit size of 39000. Its isoelectric point is at pH 6.8. It acts optimally at pH 8.6 with a Km of 2.2 mM. It does not hydrolyse L-glutamine and its reaction is inhibited competitively by D-aspartic acid and D-asparagine as well as by Ir asparagine analogues with substituents at the 0 position.

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Authors and Affiliations

  1. Laboratory of Biochemistry, Faculty of Chemistry, Aristotelian University of Thessaloniki, Thessaloniki, Greece

    D. J. Triantafillou, J. G. Georgatsos & D. A. Kyriakidis

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  1. D. J. Triantafillou
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  2. J. G. Georgatsos
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  3. D. A. Kyriakidis
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Triantafillou, D.J., Georgatsos, J.G. & Kyriakidis, D.A. Purification and properties of a membrane-bound L-asparaginase of Tetrahymena pyriformis . Mol Cell Biochem 81, 43–51 (1988). https://doi.org/10.1007/BF00225652

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  • DOI: https://doi.org/10.1007/BF00225652

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