| Functional analysis, overexpression, and kinetic characterization of pyruvate kinase from Plasmodium falciparum. | |
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MedLine Citation:
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PMID: 15567170 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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The important role of pyruvate kinase during malarial infection has prompted the cloning of a cDNA encoding Plasmodium falciparum pyruvate kinase (pfPyrK), using mRNA from intraerythrocytic-stage malaria parasites. The full-length cDNA encodes a protein with a computed molecular weight of 55.6 kDa and an isoelectric point of 7.5. The purified recombinant pfPyrK is enzymatically active and exists as a homotetramer in its active form. The enzyme exhibits hyperbolic kinetics with respect to phosphoenolpyruvate and ADP, with K(m) of 0.19 and 0.12 mM, respectively. pfPyrK is not affected by fructose-1,6-bisphosphate, a general activating factor of pyruvate kinase for most species. Glucose-6-phosphate, an activator of the Toxoplasma gondii enzyme, does not affect pfPyrK activity. Similar to rabbit pyruvate kinase, pfPyrK is susceptible to inactivation by 1mM pyridoxal-5'-phosphate, but to a lesser extent. A screen for inhibitors to pfPyrK revealed that it is markedly inhibited by ATP and citrate. Detailed kinetic analysis revealed a transition from hyperbolic to sigmoidal kinetics for PEP in the presence of citrate, as well as competitive inhibitory behavior for ATP with respect to PEP. Citrate exhibits non-competitive inhibition with respect to ADP with a K(i) of 0.8mM. In conclusion, P. falciparum expresses an active pyruvate kinase during the intraerythrocytic-stage of its developmental cycle that may play important metabolic roles during infection. |
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Authors:
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Maurice Chan; Tiow-Suan Sim |
Publication Detail:
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Type: Journal Article; Research Support, Non-U.S. Gov't |
Journal Detail:
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Title: Biochemical and biophysical research communications Volume: 326 ISSN: 0006-291X ISO Abbreviation: Biochem. Biophys. Res. Commun. Publication Date: 2005 Jan |
Date Detail:
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Created Date: 2004-11-29 Completed Date: 2005-02-03 Revised Date: 2006-11-15 |
Medline Journal Info:
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Nlm Unique ID: 0372516 Medline TA: Biochem Biophys Res Commun Country: United States |
Other Details:
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Languages: eng Pagination: 188-96 Citation Subset: IM |
Affiliation:
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Department of Microbiology, Faculty of Medicine, National University of Singapore, Singapore. |
Export Citation:
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| MeSH Terms | |
Descriptor/Qualifier:
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Amino Acid Sequence Animals Cells, Cultured Cloning, Molecular Conserved Sequence Enzyme Activation Escherichia coli / enzymology, genetics Evolution, Molecular Kinetics Molecular Sequence Data Plasmodium falciparum / enzymology*, genetics Pyruvate Kinase / chemistry*, classification, genetics Recombinant Proteins / biosynthesis, chemistry Sequence Analysis, Protein* Sequence Homology, Amino Acid Species Specificity Structure-Activity Relationship Substrate Specificity |
| Chemical | |
Reg. No./Substance:
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0/Recombinant Proteins; EC 2.7.1.40/Pyruvate Kinase |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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