Mapping the proteome of Leishmania Viannia parasites using two-dimensional polyacrylamide gel electrophoresis and associated technologies
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Instituto Nacional de Salud
2003
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| _version_ | 1876426813598597120 |
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| author | John Walker |
| author_facet | John Walker |
| contents | Mapping the proteome of Leishmania Viannia parasites using two-dimensional polyacrylamide gel electrophoresis and associated technologies John Walker Rafael Góngora Nathalie Acestor Manfredo Quadroni Nicolas Fasel Nancy G. Saravia Medicina two proteome proteomics Leishmania dimensional polyacrylamide gel electrophoresis In this study we have demonstrated the potential of two-dimensional electrophoresis (2DE)-based technologies as tools for characterization of the Leishmania proteome (the expressedprotein complement of the genome). Standardized neutral range (pH 5-7) proteome maps ofLeishmania (Viannia) guyanensis and Leishmania (Viannia) panamensis promastigotes werereproducibly generated by 2DE of soluble parasite extracts, which were prepared using lysisbuffer containing urea and nonidet P-40 detergent. The Coomassie blue and silver nitratestaining systems both yielded good resolution and representation of protein spots, enablingthe detection of approximately 800 and 1,500 distinct proteins, respectively. Several referenceprotein spots common to the proteomes of all parasite species/strains studied were isolatedand identified by peptide mass spectrometry (LC-ES-MS/MS), and bioinformatics approachesas members of the heat shock protein family, ribosomal protein S12, kinetoplast membraneprotein 11 and a hypothetical Leishmania-specific 13 kDa protein of unknown function.Immunoblotting of Leishmania protein maps using a monoclonal antibody resulted in the specificdetection of the 81.4 kDa and 77.5 kDa subunits of paraflagellar rod proteins 1 and 2, respectively.Moreover, differences in protein expression profiles between distinct parasite clones werereproducibly detected through comparative proteome analyses of paired maps using imageanalysis software. These data illustrate the resolving power of 2DE-based proteome analysis.The production and basic characterization of good quality Leishmania proteome maps providesan essential first step towards comparative protein expression studies aimed at identifying themolecular determinants of parasite drug resistance and virulence, as well as discovering newdrug and vaccine targets 2003 artículo científico 0120-4157 https://www.redalyc.org/articulo.oa?id=84323205 en http://www.redalyc.org/revista.oa?id=843 Biomédica application/pdf Instituto Nacional de Salud Biomédica (Colombia) Num.2 Vol.23 |
| format | Artículo científico |
| id | redalyc_84323205 |
| institution | Redalyc |
| language | en |
| publishDate | 2003 |
| publisher | Instituto Nacional de Salud |
| spellingShingle | Mapping the proteome of Leishmania Viannia parasites using two-dimensional polyacrylamide gel electrophoresis and associated technologies John Walker Medicina two proteome proteomics Leishmania dimensional polyacrylamide gel electrophoresis Mapping the proteome of Leishmania Viannia parasites using two-dimensional polyacrylamide gel electrophoresis and associated technologies John Walker Rafael Góngora Nathalie Acestor Manfredo Quadroni Nicolas Fasel Nancy G. Saravia Medicina two proteome proteomics Leishmania dimensional polyacrylamide gel electrophoresis In this study we have demonstrated the potential of two-dimensional electrophoresis (2DE)-based technologies as tools for characterization of the Leishmania proteome (the expressedprotein complement of the genome). Standardized neutral range (pH 5-7) proteome maps ofLeishmania (Viannia) guyanensis and Leishmania (Viannia) panamensis promastigotes werereproducibly generated by 2DE of soluble parasite extracts, which were prepared using lysisbuffer containing urea and nonidet P-40 detergent. The Coomassie blue and silver nitratestaining systems both yielded good resolution and representation of protein spots, enablingthe detection of approximately 800 and 1,500 distinct proteins, respectively. Several referenceprotein spots common to the proteomes of all parasite species/strains studied were isolatedand identified by peptide mass spectrometry (LC-ES-MS/MS), and bioinformatics approachesas members of the heat shock protein family, ribosomal protein S12, kinetoplast membraneprotein 11 and a hypothetical Leishmania-specific 13 kDa protein of unknown function.Immunoblotting of Leishmania protein maps using a monoclonal antibody resulted in the specificdetection of the 81.4 kDa and 77.5 kDa subunits of paraflagellar rod proteins 1 and 2, respectively.Moreover, differences in protein expression profiles between distinct parasite clones werereproducibly detected through comparative proteome analyses of paired maps using imageanalysis software. These data illustrate the resolving power of 2DE-based proteome analysis.The production and basic characterization of good quality Leishmania proteome maps providesan essential first step towards comparative protein expression studies aimed at identifying themolecular determinants of parasite drug resistance and virulence, as well as discovering newdrug and vaccine targets 2003 artículo científico 0120-4157 https://www.redalyc.org/articulo.oa?id=84323205 en http://www.redalyc.org/revista.oa?id=843 Biomédica application/pdf Instituto Nacional de Salud Biomédica (Colombia) Num.2 Vol.23 |
| title | Mapping the proteome of Leishmania Viannia parasites using two-dimensional polyacrylamide gel electrophoresis and associated technologies |
| topic | Medicina two proteome proteomics Leishmania dimensional polyacrylamide gel electrophoresis |
| url | https://www.redalyc.org/articulo.oa?id=84323205 |