Cloning, heterologous expression, and substrate specificities of protein farnesyltransferases from Trypanosoma cruzi and Leishmania major
| Autor: | Peter J. Myler, Kohei Yokoyama, Emily C. Speelmon, Richard T. Eastman, Wesley C. Van Voorhis, José L. Nepomuceno-Silva, Frederick S. Buckner |
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| Rok vydání: | 2002 |
| Předmět: |
Trypanosoma cruzi
Farnesyltransferase Molecular Sequence Data Antiprotozoal Agents Sf9 Spodoptera Trypanosoma brucei Substrate Specificity Mice parasitic diseases Animals Leishmania major Amino Acid Sequence Cloning Molecular Molecular Biology Gene Mice Inbred BALB C Alkyl and Aryl Transferases biology Sequence Analysis DNA DNA Protozoan biology.organism_classification Molecular biology Recombinant Proteins Rats Biochemistry Drug Design biology.protein Protein prenylation Parasitology Heterologous expression Baculoviridae |
| Zdroj: | Molecular and Biochemical Parasitology. 122:181-188 |
| ISSN: | 0166-6851 |
| Popis: | Chagas disease and leishmaniasis are tropical diseases caused by the protozoan parasites, Trypanosoma cruzi and Leishmania species, respectively. Protein farnesyltransferase (PFT) is being investigated as a target for anti-trypanosomatid agents because inhibitors of this enzyme are highly toxic to these parasites compared to mammalian cells. Here, we report the cloning of the α- and β-subunit genes of PFT from T. cruzi and Leishmania major . The proteins encoded by these genes are considerably larger than those of mammalian PFTs due to the presence of a number of inserts of >25 amino acids that map to junctions between helical structural elements. These inserts are not part of the active site or the interface between the two subunits. Northern blots demonstrate expression of messenger RNA for the PFT subunits in both mammalian and insect life-cycle stages of these parasites. The T. cruzi , Trypanosoma brucei , and L. major PFTs were overexpressed in the Sf9 cell/baculovirus system as active enzyme forms. Kinetic studies with a panel of CALX-containing peptides with all 20 amino acids in the X-position show that trypanosomatid PFTs have similar substrate specificities and these are different from the mammalian PFT substrate specificity patterns. |
| Databáze: | OpenAIRE |
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