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The Craft of Peroxisome Purification—A Technical Survey Through the Decades

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Proteomics of Peroxisomes

Part of the book series: Subcellular Biochemistry ((SCBI,volume 89))

Abstract

Purification technologies are one of the working horses in organelle proteomics studies as they guarantee the separation of organelle-specific proteins from the background contamination by other subcellular compartments. The development of methods for the separation of organelles was a major prerequisite for the initial detection and characterization of peroxisome as a discrete entity of the cell. Since then, isolated peroxisomes fractions have been used in numerous studies in order to characterize organelle-specific enzyme functions, to allocate the peroxisome-specific proteome or to unravel the organellar membrane composition. This review will give an overview of the fractionation methods used for the isolation of peroxisomes from animals, plants and fungi. In addition to “classic” centrifugation-based isolation methods, relying on the different densities of individual organelles, the review will also summarize work on alternative technologies like free-flow-electrophoresis or flow field fractionation which are based on distinct physicochemical parameters. A final chapter will further describe how different separation methods and quantitative mass spectrometry have been used in proteomics studies to assign the proteome of PO.

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Abbreviations

Tris-HCl:

Tris (hydroxymethyl) aminomethane hydrochloride

MOPS:

4-Morpholinepropane sulfonic acid

Tricine:

N-[Tris(hydroxymethyl)methyl]glycine

EDTA:

Ethylenediaminetetraacetic acid

PMSF:

Phenylmethanesulfonyl fluoride,

DTT:

1,4-Dithiothreitol

LM fraction:

Light mitochondrial fraction

HM fraction:

Heavy mitochondrial fraction

PTS:

Peroxisome targeting sequence

FFE:

Free flow electrophoresis

FlFFF:

Flow field-flow fractionation

DIGE:

Difference Gel Electrophoresis

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Acknowledgements

We apologize to those whose work has not been cited owing to space limitations. We are grateful to Profs. L. A. del Río and F. J. Corpas for providing images from plant Percoll and sucrose gradients shown in Fig. 1. We thank the FFE Service GmbH, Feldkirchen, Germany for the FFE illustrations presented in Fig. 3. This work was supported by the MEAMEDMA Anschubförderung of the Medical Faculty Mannheim, University of Heidelberg and by the German Research Foundation (DFG 397476530).

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  1. Institute for Neuroanatomy, Centre for Biomedicine and Medical Technology Mannheim, Medical Faculty Mannheim, University of Heidelberg, Heidelberg, Germany

    Markus Islinger & Andreas Manner

  2. Department of Medical Cell Biology, Institute of Anatomy, University of Heidelberg, Heidelberg, Germany

    Alfred Völkl

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  1. Markus Islinger
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  2. Andreas Manner
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Correspondence to Markus Islinger .

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  1. Department of Biochemistry, Cell and Molecular Biology of Plants, Estación Experimental del Zaidín, CSIC, Granada, Spain

    Luis A. del Río

  2. Department of Biosciences, University of Exeter, Exeter, UK

    Michael Schrader

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Islinger, M., Manner, A., Völkl, A. (2018). The Craft of Peroxisome Purification—A Technical Survey Through the Decades. In: del Río, L., Schrader, M. (eds) Proteomics of Peroxisomes. Subcellular Biochemistry, vol 89. Springer, Singapore. https://doi.org/10.1007/978-981-13-2233-4_4

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