Cell Fractionation

  • Melissa Petiti
  • Laetitia Houot
  • Denis Duché
Part of the Methods in Molecular Biology book series (MIMB, volume 1615)


Protein function is generally dependent on its subcellular localisation. In Gram-negative bacteria such as Escherichia coli, a protein can be targeted to five different compartments: the cytoplasm, the inner membrane, the periplasm, the outer membrane and the extracellular medium. Different approaches can be used to determine the protein localisation within a cell such as in silico identification of protein signal sequences and motifs, electron microscopy and immunogold labelling, optical fluorescence microscopy, and biochemical technics. In this chapter, we describe a simple and efficient method to isolate the different compartments of Escherichia coli by a fractionation method and to determine the presence of the protein of interest. For inner membrane proteins we propose a method to discriminate between integral and peripheral membrane proteins.

Key words

Spheroplast Peptidoglycan Osmotic shock Freeze and thaw Protein solubilisation Membrane Subcellular localisation 



This work was supported by the Centre National de la Recherche Scientifique and the Agence National de la Recherche (ANR-14-CE09-0023).


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© Springer Science+Business Media LLC 2017

Authors and Affiliations

  1. 1.Laboratoire d’Ingénierie des Systèmes Macromoléculaires (LISM, UMR 7255)Institut de Microbiologie de la Méditerranée (IMM), Aix-Marseille Université—Centre National de la Recherche Scientifique (CNRS)Marseille Cedex 20France

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