Towards the recovery of hydrophobic proteins on two-dimensional electrophoresis gels
| Autor: | Patrick Doumas, Véronique Santoni, Jérôme Garin, Michel Rossignol, Thierry Rabilloud, David Rouquié, Monique Mansion, Sylvie Kieffer |
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| Přispěvatelé: | Biochimie et Physiologie Moléculaire des Plantes (BPMP), Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS), Plateforme de Spectrométrie de Masse Protéomique - Mass Spectrometry Proteomics Platform (MSPP), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA), Equipe Intégration des signalisations nutritionnelles (INTEGRATION), Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), Institut National de la Recherche Agronomique (INRA)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS) |
| Jazyk: | angličtina |
| Rok vydání: | 1999 |
| Předmět: |
purification
Octoxynol [SDV]Life Sciences [q-bio] Detergents Clinical Biochemistry Acrylic Resins Arabidopsis Carbonates membrane plasmique Cell Fractionation plasma membrane Biochemistry Polyethylene Glycols Analytical Chemistry kjeldahl method Chaps Electrophoresis Gel Two-Dimensional Integral membrane protein protéome Plant Proteins hydrophobicity Chromatography solubilizing BIOCHIMIE Chemistry refining Peripheral membrane protein arabidopsis thaliana Cholic Acids two-dimensional electrophoresis hydrophobicité Plant Leaves Electrophoresis Membrane électrophorèse bidimensionnelle Solubility Membrane protein protéine solubilisation Proteome technique analytique Immobilized pH gradient protein |
| Zdroj: | Electrophoresis Electrophoresis, Wiley-VCH Verlag, 1999, 20, pp.705-711. ⟨10.1002/(SICI)1522-2683(19990101)20:4/53.0.CO;2-Q⟩ From Genome to Proteome: Advances in the Practice and Application of Proteomics |
| ISSN: | 0173-0835 1522-2683 |
| Popis: | An extensive proteomic approach relies on the possibility to visualize and analyze various types of proteins, including hydrophobic proteins which are rarely detectable on two-dimensional electrophoresis (2-DE) gels. In this study, two methods were employed for the purification of hydrophobic proteins from Arabidopsis thaliana leaf plasma membrane (PM) model plants, prior to analysis on 2-DE immobilized pH gradient (IPG) gels. Solubilization efficiency of two detergents, (3-[(3-cholomidopropyl)-1-propanesulfonic acid (CHAPS) and C8phi, were tested for the recovery of hydrophobic proteins. An immunological approach was used to determine the efficiency of the above methods. Fractionation of proteins by Triton X-114 combined with solubilization with CHAPS resulted in the inability to detect hydrophobic proteins on 2-DE gels. The use of C8phi for protein solubilization did not improve this result. On the contrary, after treatment of membranes with alkaline buffer, the solubilization of PM proteins with detergent C8phi permitted the recovery of such proteins on 2-DE gels. The combination of membrane washing and the use of zwitterionic detergent resulted in the resolution of several integral proteins and the disappearance of peripheral proteins. In the resolution of expressed genome proteins, both large pH gradients in the first dimension and various acrylamide concentrations in the second dimension must be used. Notwithstanding, it is important to combine various sample treatments and different detergents in order to resolve soluble and hydrophobic proteins. |
| Databáze: | OpenAIRE |
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