Comparison of methods for simultaneous identification of bacterial species and determination of metabolic activity by protein‐based stable isotope probing (Protein‐SIP) experiments

Jehmlich, Nico; Schmidt, Frank; Taubert, Martin; Seifert, Jana; Bergen, Martin von; Richnow, Hans H.; Vogt, Carsten

doi:10.1002/rcm.4084

Cited by 29 publications

(28 citation statements)

References 43 publications

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“…However, gradient centrifugation can only resolve nucleic acids with large differences in the degree of label incorporation. Recently a protein-based SIP method was developed that uses mass spectrometry (MS) to determine the extent of 13 C incorporation of peptides and proteins (4–9). Because the degree of label incorporation in proteins and peptides can be determined to high resolution by mass spectrometry, and as proteins and peptides contain sequence information that links each molecule to its organism of origin, protein-based SIP methods enable determination of low levels of isotope incorporation into microorganisms that can be resolved at the strain level.…”

mentioning

confidence: 99%

Quantitative Tracking of Isotope Flows in Proteomes of Microbial Communities

Pan

Fischer

Hyatt

et al. 2011

Molecular & Cellular Proteomics

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Stable isotope probing (SIP) has been used to track nutrient flows in microbial communities, but existing protein-based SIP methods capable of quantifying the degree of label incorporation into peptides and proteins have been demonstrated only by targeting usually less than 100 proteins per sample. Our method automatically (i) identifies the sequence of and (ii) quantifies the degree of heavy atom enrichment for thousands of proteins from microbial community proteome samples. These features make our method suitable for comparing isotopic differences between closely related protein sequences, and for detecting labeling patterns in low-abundance proteins or proteins derived from rare community members. The proteomic SIP method was validated using proteome samples of known stable isotope incorporation levels at 0.4%, ∼50%, and ∼98%. The method was then used to monitor incorporation of 15N into established and regrowing microbial biofilms. The results indicate organism-specific migration patterns from established communities into regrowing communities and provide insights into metabolism during biofilm formation. The proteomic SIP method can be extended to many systems to track fluxes of 13C or 15N in microbial communities.

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mentioning

confidence: 99%

Quantitative Tracking of Isotope Flows in Proteomes of Microbial Communities

Pan

Fischer

Hyatt

et al. 2011

Molecular & Cellular Proteomics

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“…According to the authors of this method (Jehmlich et al 2009) this can be overcome by analysing at least three peptides per protein in each case. However, this can result in a very time consuming analysis.…”

Section: Protein-stable Isotope Probing (Protein-sip)mentioning

confidence: 99%

An appraisal of methods for linking environmental processes to specific microbial taxa

Gutierrez-Zamora

Manefield

2010

Rev Environ Sci Biotechnol

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The last decade has witnessed a revolution in the development of methods and technology available to investigate the ecological roles of microorganisms in the environment. As a consequence, microbial ecologists have gained a better understanding of the functional aspects of microorganisms in marine, groundwater and freshwater systems, soils, sediments, hot springs, wastewater treatment plants, landfills, the rhizosphere and the animal gut. This review provides a compilation and critical comparison of the currently available methods linking microbial function with phylogeny, including a description and advantages and limitations of each method. Examples are also provided to illustrate their application. The ongoing improvements of these 'function-identity' methods points to a bright future in our understanding of complex ecological processes and to improved management of microbe dependent ecosystem services.

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“…DNA-based SIP will measure primarily newly formed cells, while RNA-based SIP will address also non-growing microorganisms and is highly dynamic, especially mRNA (Dumont et al, 2011). Jehmlich et al (Jehmlich et al, 2009) developed a concept for analyzing carbon and nitrogen fluxes in microbial communities by employing protein-based SIP in metabolic labeling experiments with stable isotope labeled substrates.…”

Section: What Is Needed To Mechanistically Model Complex Communities:mentioning

confidence: 99%

Toward quantitative understanding on microbial community structure and functioning: a modeling-centered approach using degradation of marine oil spills as example

Röling¹,

Bodegom

2014

Front. Microbiol.

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Molecular ecology approaches are rapidly advancing our insights into the microorganisms involved in the degradation of marine oil spills and their metabolic potentials. Yet, many questions remain open: how do oil-degrading microbial communities assemble in terms of functional diversity, species abundances and organization and what are the drivers? How do the functional properties of microorganisms scale to processes at the ecosystem level? How does mass flow among species, and which factors and species control and regulate fluxes, stability and other ecosystem functions? Can generic rules on oil-degradation be derived, and what drivers underlie these rules? How can we engineer oil-degrading microbial communities such that toxic polycyclic aromatic hydrocarbons are degraded faster? These types of questions apply to the field of microbial ecology in general. We outline how recent advances in single-species systems biology might be extended to help answer these questions. We argue that bottom-up mechanistic modeling allows deciphering the respective roles and interactions among microorganisms. In particular constraint-based, metagenome-derived community-scale flux balance analysis appears suited for this goal as it allows calculating degradation-related fluxes based on physiological constraints and growth strategies, without needing detailed kinetic information. We subsequently discuss what is required to make these approaches successful, and identify a need to better understand microbial physiology in order to advance microbial ecology. We advocate the development of databases containing microbial physiological data. Answering the posed questions is far from trivial. Oil-degrading communities are, however, an attractive setting to start testing systems biology-derived models and hypotheses as they are relatively simple in diversity and key activities, with several key players being isolated and a high availability of experimental data and approaches.

show abstract

Comparison of methods for simultaneous identification of bacterial species and determination of metabolic activity by protein‐based stable isotope probing (Protein‐SIP) experiments

Cited by 29 publications

References 43 publications

Quantitative Tracking of Isotope Flows in Proteomes of Microbial Communities

Quantitative Tracking of Isotope Flows in Proteomes of Microbial Communities

An appraisal of methods for linking environmental processes to specific microbial taxa

Toward quantitative understanding on microbial community structure and functioning: a modeling-centered approach using degradation of marine oil spills as example

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