A protein-centric approach for the identification of folate enzymes from the malarial parasite, Plasmodium falciparum , using OFFGEL™ solution-based isoelectric focussing and mass spectrometry
Abstract Background Plasmodium species are difficult to study using proteomic technology because they contain large amounts of haemoglobin-derived products (HDP), generated by parasite breakdown of host haemoglobin. HDP are known to interfere with isoelectric focussing, a cornerstone of fractionatio...
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ftdoajarticles:oai:doaj.org/article:f0b4dd8e251e45f3b5ab3c5f4722cf8b 2023-05-15T15:08:22+02:00 A protein-centric approach for the identification of folate enzymes from the malarial parasite, Plasmodium falciparum , using OFFGEL™ solution-based isoelectric focussing and mass spectrometry Sims Paul FG Hyde John E O'Cualain Ronan DM 2010-10-01T00:00:00Z https://doi.org/10.1186/1475-2875-9-286 https://doaj.org/article/f0b4dd8e251e45f3b5ab3c5f4722cf8b EN eng BMC http://www.malariajournal.com/content/9/1/286 https://doaj.org/toc/1475-2875 doi:10.1186/1475-2875-9-286 1475-2875 https://doaj.org/article/f0b4dd8e251e45f3b5ab3c5f4722cf8b Malaria Journal, Vol 9, Iss 1, p 286 (2010) Arctic medicine. Tropical medicine RC955-962 Infectious and parasitic diseases RC109-216 article 2010 ftdoajarticles https://doi.org/10.1186/1475-2875-9-286 2022-12-30T22:27:46Z Abstract Background Plasmodium species are difficult to study using proteomic technology because they contain large amounts of haemoglobin-derived products (HDP), generated by parasite breakdown of host haemoglobin. HDP are known to interfere with isoelectric focussing, a cornerstone of fractionation strategies for the identification of proteins by mass spectrometry. In addition to the challenge presented by this material, as in most proteomes, there exists in this parasite a considerable dynamic range between proteins of high and low abundance. The enzymes of the folate pathway, a proven and widely used drug target, are included in the latter class. Methods This report describes a work-flow utilizing a parasite-specific extraction protocol that minimizes release of HDP into the lysate, followed by in-solution based OFFGEL™ electrophoresis at the protein level, trypsin digestion and mass spectrometric analysis. Results It is demonstrated that, by removing HDP from parasite lysates, OFFGEL™-mediated protein separation is able to deliver reduced complexity protein fractions. Importantly, proteins with similar and predictable physical properties are sharply focussed within such fractions. Conclusions By following this novel workflow, data have been obtained which allow the unequivocal experimental identification by mass spectrometry of four of the six proteins involved in folate biosynthesis and recycling. Article in Journal/Newspaper Arctic Directory of Open Access Journals: DOAJ Articles Arctic Malaria Journal 9 1 286 |
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Open Polar |
collection |
Directory of Open Access Journals: DOAJ Articles |
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ftdoajarticles |
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English |
topic |
Arctic medicine. Tropical medicine RC955-962 Infectious and parasitic diseases RC109-216 |
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Arctic medicine. Tropical medicine RC955-962 Infectious and parasitic diseases RC109-216 Sims Paul FG Hyde John E O'Cualain Ronan DM A protein-centric approach for the identification of folate enzymes from the malarial parasite, Plasmodium falciparum , using OFFGEL™ solution-based isoelectric focussing and mass spectrometry |
topic_facet |
Arctic medicine. Tropical medicine RC955-962 Infectious and parasitic diseases RC109-216 |
description |
Abstract Background Plasmodium species are difficult to study using proteomic technology because they contain large amounts of haemoglobin-derived products (HDP), generated by parasite breakdown of host haemoglobin. HDP are known to interfere with isoelectric focussing, a cornerstone of fractionation strategies for the identification of proteins by mass spectrometry. In addition to the challenge presented by this material, as in most proteomes, there exists in this parasite a considerable dynamic range between proteins of high and low abundance. The enzymes of the folate pathway, a proven and widely used drug target, are included in the latter class. Methods This report describes a work-flow utilizing a parasite-specific extraction protocol that minimizes release of HDP into the lysate, followed by in-solution based OFFGEL™ electrophoresis at the protein level, trypsin digestion and mass spectrometric analysis. Results It is demonstrated that, by removing HDP from parasite lysates, OFFGEL™-mediated protein separation is able to deliver reduced complexity protein fractions. Importantly, proteins with similar and predictable physical properties are sharply focussed within such fractions. Conclusions By following this novel workflow, data have been obtained which allow the unequivocal experimental identification by mass spectrometry of four of the six proteins involved in folate biosynthesis and recycling. |
format |
Article in Journal/Newspaper |
author |
Sims Paul FG Hyde John E O'Cualain Ronan DM |
author_facet |
Sims Paul FG Hyde John E O'Cualain Ronan DM |
author_sort |
Sims Paul FG |
title |
A protein-centric approach for the identification of folate enzymes from the malarial parasite, Plasmodium falciparum , using OFFGEL™ solution-based isoelectric focussing and mass spectrometry |
title_short |
A protein-centric approach for the identification of folate enzymes from the malarial parasite, Plasmodium falciparum , using OFFGEL™ solution-based isoelectric focussing and mass spectrometry |
title_full |
A protein-centric approach for the identification of folate enzymes from the malarial parasite, Plasmodium falciparum , using OFFGEL™ solution-based isoelectric focussing and mass spectrometry |
title_fullStr |
A protein-centric approach for the identification of folate enzymes from the malarial parasite, Plasmodium falciparum , using OFFGEL™ solution-based isoelectric focussing and mass spectrometry |
title_full_unstemmed |
A protein-centric approach for the identification of folate enzymes from the malarial parasite, Plasmodium falciparum , using OFFGEL™ solution-based isoelectric focussing and mass spectrometry |
title_sort |
protein-centric approach for the identification of folate enzymes from the malarial parasite, plasmodium falciparum , using offgel™ solution-based isoelectric focussing and mass spectrometry |
publisher |
BMC |
publishDate |
2010 |
url |
https://doi.org/10.1186/1475-2875-9-286 https://doaj.org/article/f0b4dd8e251e45f3b5ab3c5f4722cf8b |
geographic |
Arctic |
geographic_facet |
Arctic |
genre |
Arctic |
genre_facet |
Arctic |
op_source |
Malaria Journal, Vol 9, Iss 1, p 286 (2010) |
op_relation |
http://www.malariajournal.com/content/9/1/286 https://doaj.org/toc/1475-2875 doi:10.1186/1475-2875-9-286 1475-2875 https://doaj.org/article/f0b4dd8e251e45f3b5ab3c5f4722cf8b |
op_doi |
https://doi.org/10.1186/1475-2875-9-286 |
container_title |
Malaria Journal |
container_volume |
9 |
container_issue |
1 |
container_start_page |
286 |
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1766339742144659456 |