| Article ID | Journal | Published Year | Pages | File Type |
|---|---|---|---|---|
| 2830330 | Molecular and Biochemical Parasitology | 2006 | 6 Pages |
A full-length dihydroorotase (DHOase) sequence was cloned from a Toxoplasma gondii tachyzoite cDNA library. The sequence had a calculated molecular mass of 44.2 kDa and a pI of 5.72, and was most similar to type IIa DHOases. A recombinant protein was expressed and purified with a yield of ∼20 mg L−1 of cell culture. Polyclonal antibodies raised against purified recombinant protein reacted with a band of the expected molecular mass in tachyzoite extracts. Specific activities of 18.3 μmol/min/mg in the biosynthetic direction and 18.4 μmol/min/mg in the degradative direction, with Km, carbamyl aspartate = 323 μM and Km, dihydroorotate = 64.3 μM, were measured for purified recombinant protein. Size exclusion chromatography/laser light scattering showed a single, monodisperse peak with a molecular mass of 45.6 kDa, suggesting that the native protein is a monomer.
