Typical metaproteomics workflow ranging from microbiome sampling over MS/MS acquisition to database-driven taxonomic and functional characterization
Source: BAM, section eScience
Metaproteomics can be understood to be the large-scale profiling of the entire protein set produced by a complex microbial ecosystem such as the human gut microbiome. It can reveal the functionality of the complex microbial communities that surround us: for example, the human gut harbors many beneficial (yet sometimes harmful) bacteria, but microbial communities are also important in the context of renewable energy sources in biogas plants. The Metaproteomics Initiative is an international effort with members from Germany, Belgium, Norway, Italy, and the United States. More information on the initiative can be found here in a commentary published in the journal Microbiome.
A community-driven benchmarking study was set up, called Critical Assessment of MetaProteome Investigation (CAMPI). In this study, the involved labs were supplied with two different microbiome samples: an artificial human intestine model and a real human fecal sample. To identify and quantify proteins and taxa from these samples, the labs then applied their individual bioanalytical workflows with the steps of sample preparation, mass spectrometry analysis, and bioinformatic tools to process and analyze the generated data. By evaluating the results generated by each workflow, the involved labs assessed the effects of the workflows on peptide, protein, taxonomic and functional level.
The main result of the CAMPI study was that the evaluated workflows were very robust. In detail, the number and identities of proteins in each sample type were highly similar across the different protocols and analyses used by the individual labs. However, it was observed there was notable variability in the number of peptides identified by each workflow. These differences were linked to variation in sample preparation prior to mass spectrometry analysis. Overall, although the outcome related to protein content of the samples was similar using each workflow, the comparative study revealed that critical steps of sample preparation techniques influence the detection of certain peptides and proteins. The findings of this study serve as the basis for future multi-laboratory studies that are required to work towards consensus approaches for analytics in metaproteomics.
Critical assessment of MetaProteome Investigation (CAMPI): A multi-laboratory comparison of established workflows
T. Van Den Bossche, B. Kunath, K. Schallert ... T. Muth
published in Nature Communication, Vol. 12, article 7305, 2021
BAM section eScience