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Protocol for Performing Protein Stable Isotope Probing (Protein-SIP) Experiments

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Hydrocarbon and Lipid Microbiology Protocols

Part of the book series: Springer Protocols Handbooks ((SPH))

Abstract

Protein-based stable isotope probing (protein-SIP) was developed to link microbial-specific metabolic function to phylogenetic information. The SIP-concept is based on the application of stable isotope-labeled compounds, 13C or 15N as substrates supplied to microbial communities grown within microcosm systems. Microorganisms capable of utilizing these substrates become labeled upon metabolism and biosynthesis. The sum of all proteins represents the functional status of the living organisms, so that the degree of heavy isotope incorporation represents a proxy for substrate assimilation. The incorporation into peptides/proteins is calculated using the non-negative least squares approach. In addition, the peptide sequence can discern the active microorganisms resolved at species-level, if species specific genomic information and unique peptide information is available. The protein-SIP approach is a useful method to investigate the composition and the functional state of microbial communities. Here, we provide a guideline for incubating microbial cultures with labeled substrates and a protocol for protein-SIP. The protocol guides the reader through the proteomics pipeline, including protein extraction, protein separation, and the mass spectrometric analysis. The calculation of stable isotope incorporation into peptides/proteins will be explained in more detail.

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Author information

Authors and Affiliations

  1. Department of Molecular Systems Biology, Helmholtz Centre for Environmental Research – UFZ, Permoserstr. 15, 04318, Leipzig, Germany

    Nico Jehmlich & Martin von Bergen

  2. Department of Metabolomics, Helmholtz Centre for Environmental Research – UFZ, Permoserstr. 15, 04318, Leipzig, Germany

    Martin von Bergen

  3. Department of Chemistry and Bioscience, Aalborg University, Fredrik Bajers Vej 7, 9220, Aalborg, Denmark

    Martin von Bergen

  4. Faculty of Biosciences, Pharmacy and Psychology, Institute of Biochemistry, University Leipzig, Leipzig, Germany

    Martin von Bergen

Authors
  1. Nico Jehmlich
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  2. Martin von Bergen
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Corresponding author

Correspondence to Nico Jehmlich .

Editor information

Editors and Affiliations

  1. Dept. Biological Sciences, University of Essex Dept. Biological Sciences, Colchester, Essex, United Kingdom

    Terry J. McGenity

  2. Braunschweig, Germany

    Kenneth N. Timmis

  3. Department of Biology, University of the Balearic Islands and Mediterranean Institute for Advanced Studies (IMEDEA, UIB-CSIC), Palma de Mallorca, Spain

    Balbina Nogales

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© 2016 Springer-Verlag Berlin Heidelberg

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Jehmlich, N., von Bergen, M. (2016). Protocol for Performing Protein Stable Isotope Probing (Protein-SIP) Experiments. In: McGenity, T., Timmis, K., Nogales , B. (eds) Hydrocarbon and Lipid Microbiology Protocols. Springer Protocols Handbooks. Springer, Berlin, Heidelberg. https://doi.org/10.1007/8623_2016_209

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  • DOI: https://doi.org/10.1007/8623_2016_209

  • Published:

  • Publisher Name: Springer, Berlin, Heidelberg

  • Print ISBN: 978-3-662-50449-9

  • Online ISBN: 978-3-662-50450-5

  • eBook Packages: Springer Protocols

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