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An Integrated Workflow for Enhanced Taxonomic and Functional Coverage of the Mouse Faecal Metaproteome

Nicolas Nalpas, View ORCID ProfileLesley Hoyles, Viktoria Anselm, Tariq Ganief, Laura Martinez-Gili, Cristina Grau, Irina Droste-Borel, Laetitia Davidovic, Xavier Altafaj, Marc-Emmanuel Dumas, Boris Macek
doi: https://doi.org/10.1101/2020.11.17.386938
Nicolas Nalpas
1Proteome Center Tuebingen, University of Tuebingen, Germany
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Lesley Hoyles
2Biomolecular Medicine Section, Division of systems Medicine, Department of Metabolism, Digestion and Reproduction, Imperial College London, Sir Alexander Fleming building, London SW7 2AZ, UK
3Department of Biosciences, Nottingham Trent University, UK
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  • ORCID record for Lesley Hoyles
Viktoria Anselm
1Proteome Center Tuebingen, University of Tuebingen, Germany
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Tariq Ganief
1Proteome Center Tuebingen, University of Tuebingen, Germany
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Laura Martinez-Gili
2Biomolecular Medicine Section, Division of systems Medicine, Department of Metabolism, Digestion and Reproduction, Imperial College London, Sir Alexander Fleming building, London SW7 2AZ, UK
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Cristina Grau
4Bellvitge Biomedical Research Institute, Spain
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Irina Droste-Borel
1Proteome Center Tuebingen, University of Tuebingen, Germany
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Laetitia Davidovic
5Université Côte d’Azur, CNRS, Inserm, IPMC, France
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Xavier Altafaj
4Bellvitge Biomedical Research Institute, Spain
6Neurophysiology Unit, University of Barcelona - IDIBAPS, Spain
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Marc-Emmanuel Dumas
2Biomolecular Medicine Section, Division of systems Medicine, Department of Metabolism, Digestion and Reproduction, Imperial College London, Sir Alexander Fleming building, London SW7 2AZ, UK
7Genomic and Environmental Medicine, National Heart & Lung Institute, Faculty of Medicine, Imperial College London, London, SW3 6KY, United Kingdom
8European Genomic Institute for Diabetes, INSERM UMR 1283, CNRS UMR 8199, Institut Pasteur de Lille, Lille University Hospital, University of Lille, 59045 Lille, France
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Boris Macek
1Proteome Center Tuebingen, University of Tuebingen, Germany
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  • For correspondence: boris.macek@uni-tuebingen.de
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Abstract

The intestinal microbiota plays a key role in shaping host homeostasis by regulating metabolism, immune responses and behaviour. Its dysregulation has been associated with metabolic, immune and neuropsychiatric disorders and is accompanied by changes in bacterial metabolic regulation. Although proteomics is well suited for analysis of individual microbes, metaproteomics of faecal samples is challenging due to the physical structure of the sample, presence of contaminating host proteins and coexistence of hundreds of species. Furthermore, there is a lack of consensus regarding preparation of faecal samples, as well as downstream bioinformatic analyses following metaproteomic data acquisition. Here we assess sample preparation and data analysis strategies applied to mouse faeces in a typical LC-MS/MS metaproteomic experiment. We show that low speed centrifugation (LSC) of faecal samples leads to high protein identification rates and a balanced taxonomic representation. During database search, protein sequence databases derived from matched mouse faecal metagenomes provided up to four times more MS/MS identifications compared to other database construction strategies, while a two-step database search strategy led to accumulation of false positive protein identifications. Comparison of matching metaproteome and metagenome data revealed a positive correlation between protein and gene abundances, as well as significant overlap and correlation in taxonomic representation. Notably, nearly all functional categories of detected protein groups were differentially abundant in the metaproteome compared to what would be expected from the metagenome, highlighting the need to perform metaproteomics when studying complex microbiome samples.

Competing Interest Statement

The authors have declared no competing interest.

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The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC 4.0 International license.
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Posted November 17, 2020.
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An Integrated Workflow for Enhanced Taxonomic and Functional Coverage of the Mouse Faecal Metaproteome
Nicolas Nalpas, Lesley Hoyles, Viktoria Anselm, Tariq Ganief, Laura Martinez-Gili, Cristina Grau, Irina Droste-Borel, Laetitia Davidovic, Xavier Altafaj, Marc-Emmanuel Dumas, Boris Macek
bioRxiv 2020.11.17.386938; doi: https://doi.org/10.1101/2020.11.17.386938
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An Integrated Workflow for Enhanced Taxonomic and Functional Coverage of the Mouse Faecal Metaproteome
Nicolas Nalpas, Lesley Hoyles, Viktoria Anselm, Tariq Ganief, Laura Martinez-Gili, Cristina Grau, Irina Droste-Borel, Laetitia Davidovic, Xavier Altafaj, Marc-Emmanuel Dumas, Boris Macek
bioRxiv 2020.11.17.386938; doi: https://doi.org/10.1101/2020.11.17.386938

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