Abstract:
:The pyrimidine biosynthesis pathway in the protozoan pathogen Toxoplasma gondii is essential for parasite growth during infection. To investigate the properties of dihydroorotate dehydrogenase (TgDHOD), the fourth enzyme in the T. gondii pyrimidine pathway, we expressed and purified recombinant TgDHOD. TgDHOD exhibited a specific activity of 84U/mg, a k(cat) of 89s(-1), a K(m)=60μM for l-dihydroorotate, and a K(m)=29μM for decylubiquinone (Q(D)). Quinones lacking or having short isoprenoid side chains yielded lower k(cat)s than Q(D). As expected, fumarate was a poor electron acceptor for this family 2 DHOD. The IC(50)s determined for A77-1726, the active derivative of the human DHOD inhibitor leflunomide, and related compounds MD249 and MD209 were, 91μM, 96μM, and 60μM, respectively. The enzyme was not significantly affected by brequinar or TTFA, known inhibitors of human DHOD, or by atovaquone. DSM190, a known inhibitor of Plasmodium falciparum DHOD, was a poor inhibitor of TgDHOD. TgDHOD exhibits a lengthy 157-residue N-terminal extension, consistent with a potential organellar targeting signal. We constructed C-terminally c-myc tagged TgDHODs to examine subcellular localization of TgDHOD in transgenic parasites expressing the tagged protein. Using both exogenous and endogenous expression strategies, anti-myc fluorescence signal colocalized with antibodies against the mitochondrial marker ATPase. These findings demonstrate that TgDHOD is associated with the parasite's mitochondrion, revealing this organelle as the site of orotate production in T. gondii. The TgDHOD gene appears to be essential because while gene tagging was successful at the TgDHOD gene locus, attempts to delete the TgDHOD gene were not successful in the KU80 background. Collectively, our study suggests that TgDHOD is an excellent target for the development of anti-Toxoplasma drugs.
journal_name
Mol Biochem Parasitoljournal_title
Molecular and biochemical parasitologyauthors
Hortua Triana MA,Huynh MH,Garavito MF,Fox BA,Bzik DJ,Carruthers VB,Löffler M,Zimmermann BHdoi
10.1016/j.molbiopara.2012.04.009subject
Has Abstractpub_date
2012-08-01 00:00:00pages
71-81issue
2eissn
0166-6851issn
1872-9428pii
S0166-6851(12)00102-8journal_volume
184pub_type
杂志文章abstract::The anaerobic, parasitic protist, Trichomonas vaginalis, is characterized by the absence of mitochondria and the presence of double membrane bound organelles called hydrogenosomes. Succinyl-coenzyme A synthetase is a hydrogenosomal enzyme which catalyzes the formation of ATP via substrate-level phosphorylation. We hav...
journal_title:Molecular and biochemical parasitology
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abstract::Hookworm disease is a debilitating worm infection that affects hundreds of millions of people. Despite the existence of anthelmintic drugs, reports have testified of a decrease in efficacy of these drugs. Therefore, it is imperative to find new drugs and drug targets for hookworm disease treatment. In this study we id...
journal_title:Molecular and biochemical parasitology
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journal_title:Molecular and biochemical parasitology
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journal_title:Molecular and biochemical parasitology
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journal_title:Molecular and biochemical parasitology
pub_type: 杂志文章
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pub_type: 杂志文章
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journal_title:Molecular and biochemical parasitology
pub_type: 杂志文章
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journal_title:Molecular and biochemical parasitology
pub_type: 杂志文章
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journal_title:Molecular and biochemical parasitology
pub_type: 杂志文章
doi:10.1016/j.molbiopara.2007.07.010
更新日期:2007-11-01 00:00:00
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journal_title:Molecular and biochemical parasitology
pub_type: 杂志文章
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更新日期:2006-07-01 00:00:00
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pub_type: 杂志文章
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pub_type: 杂志文章
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更新日期:2016-01-01 00:00:00
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journal_title:Molecular and biochemical parasitology
pub_type: 杂志文章
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journal_title:Molecular and biochemical parasitology
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journal_title:Molecular and biochemical parasitology
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journal_title:Molecular and biochemical parasitology
pub_type: 杂志文章
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journal_title:Molecular and biochemical parasitology
pub_type: 杂志文章
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更新日期:1996-12-02 00:00:00
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journal_title:Molecular and biochemical parasitology
pub_type: 杂志文章
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journal_title:Molecular and biochemical parasitology
pub_type: 杂志文章
doi:10.1016/s0166-6851(98)00066-8
更新日期:1998-08-01 00:00:00
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pub_type: 杂志文章
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更新日期:1990-03-01 00:00:00
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pub_type: 杂志文章
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journal_title:Molecular and biochemical parasitology
pub_type: 杂志文章
doi:10.1016/s0166-6851(98)00119-4
更新日期:1998-10-30 00:00:00
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journal_title:Molecular and biochemical parasitology
pub_type: 杂志文章
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journal_title:Molecular and biochemical parasitology
pub_type: 杂志文章
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更新日期:2012-07-01 00:00:00
abstract::Phosphatidylcholine (PC) is the most abundant phospholipid in the membranes of the human parasite Leishmania. It is synthesized via two metabolic routes, the de novo pathway that starts with the uptake of choline, and the threefold methylation of phosphatidylethanolamine. Choline was shown to be dispensable for Leishm...
journal_title:Molecular and biochemical parasitology
pub_type: 杂志文章
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更新日期:2014-09-01 00:00:00