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Eukaryotic Cell, December 2006, p. 2072-2078, Vol. 5, No. 12
1535-9778/06/$08.00+0     doi:10.1128/EC.00249-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Identification and Biochemical Characterization of Serine Hydroxymethyl Transferase in the Hydrogenosome of Trichomonas vaginalis ^,

Department of Microbiology, Immunology, and Molecular Genetics, University of California, Los Angeles, 609 Charles E. Young Drive East, Los Angeles, California 90095-1489,¹ Department of Chemistry and Biochemistry, University of California, Los Angeles, California 90095²

Received 4 August 2006/ Accepted 7 September 2006

Serine hydroxymethyl transferase (SHMT) is a pyridoxal phosphate (PLP)-dependent enzyme that catalyzes the reversible conversion of serine and tetrahydrofolate to glycine and methylenetetrahydrofolate. We have identified a single gene encoding SHMT in the genome of Trichomonas vaginalis, an amitochondriate, deep-branching unicellular protist. The protein possesses a putative N-terminal hydrogenosomal presequence and was shown to localize to hydrogensomes by immunofluorescence analysis, providing evidence of amino acid metabolism in this unusual organelle. In contrast to the tetrameric SHMT that exists in the mammalian host, we found that the T. vaginalis SHMT is a homodimer, as found in prokaryotes. All examined SHMT contain an 8-amino-acid conserved sequence, VTTTTHKT, containing the active-site lysyl residue (Lys 251 in TvSHMT) that forms an internal aldimine with PLP. We mutated this Lys residue to Arg and Gln and examined structural and catalytic properties of the wild-type and mutant enzymes in comparison to that reported for the mammalian protein. The oligomeric structure of the mutant K251R and K251Q TvSHMT was not affected, in contrast to that observed for comparable mutations in the mammalian enzyme. Likewise, contrary to that observed for mammalian SHMT, the catalytic activity of K251R TvSHMT was unaffected in the presence of PLP. The K251Q TvSHMT, however, was found to be inactive. These studies indicate that the active site of the parasite enzyme is distinct from its prokaryotic and eukaryotic counterparts and identify TvSHMT as a potential drug target.

Published ahead of print on 15 September 2006.

Supplemental material for this article may be found at http://ec.asm.org/.