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Exp Parasitol. 1998 Oct;90(2):175-80.

Trichomonas vaginalis: expression and characterisation of recombinant S-adenosylhomocysteinase.

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  • 1School of Biochemistry and Molecular Genetics, University of New South Wales, Sydney, 2052, Australia.


The gene encoding S-adenosylhomocysteinase activity (S-adenosylhomocysteine hydrolase, SAHH; EC in Trichomonas vaginalis has been expressed in Escherichia coli to facilitate the characterisation of the enzyme. Expression of this gene using the pQE-30 (6xHis N-terminal tag) expression system (QIAGEN) has enabled the one-step purification of 6 mg of active recombinant enzyme from a 100-ml bacterial culture by affinity chromatography using a nickel-NTA matrix. The recombinant enzyme has a molecular weight of approximately 56,000 and identification of tryptic peptides by matrix-assisted laser desorption ionisation (MALDI) mass spectrometry has shown that the purified recombinant protein is identical in primary structure to the predicted sequence. The presence of the N-terminal 6xHis tag in the recombinant enzyme did not appear to affect its kinetic and other properties, which are similar to those exhibited by the "native" enzyme present in cell-free extracts of T. vaginalis. These properties include a similar apparent Km for adenosine (20-25 microM for the recombinant and 5-10 microM for the native enzymes, respectively) and similar inhibition/inactivation patterns exhibited by adenosine analogues such as arabinosyl adenine (ara-A).

Copyright 1998 Academic Press.

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