Abstract
Protein recovery is crucial for shotgun metaproteomics to study the in situ functionality of microbial populations from complex biofilms but still poorly addressed by far. To fill this knowledge gap, we systematically evaluated the sample preparation with extraction buffers comprising four detergents for the metaproteomics analysis of a terephthalate-degrading methanogenic biofilm using an on-line two-dimensional liquid chromatography tandem mass spectrometry (2D-LC-MS/MS) system. Totally, 1018 non-repeated proteins were identified with the four treatments. On the whole, each treatment could recover the biofilm proteins with specific distributions of molecular weight, hydrophobicity, and isoelectric point. The extraction buffers containing zwitterionic and anionic detergents were found to harvest the proteins with better efficiency and quality, allowing identification up to 76.2% of total identified proteins with the LC-MS/MS analysis. According to the annotation with a relevant metagenomic database, we further observed different taxonomic profiles of bacterial and archaeal members and discriminable patterns of the functional expression among the extraction buffers used. Overall, the finding of the present study provides first insight to the effect of the detergents on the characteristics of extractable proteins from biofilm and the developed protocol combined with nano 2D-LC/MS/MS analysis can improve the metaproteomics studies on microbial functionality of biofilms in the wastewater treatment systems.
Highlights
Biofilms are the structured consortia of trophic groups of microorganisms grown in a close approximation on the surface of supporting media, which can be involved an important biological process for water/wastewater purification, bioenergy recovery, and environmental remediation
We used four different buffers in the trichloroacetic acid (TCA)-based extraction protocol that is commonly used for studies of shotgun metaproteomics [11] to recover total proteins from the methane-producing biofilm grown on the ceramic rings from an anaerobic fixed-film reactor fed with terephthalate as the sole substrate at 50 °C [4]
We evaluated the characteristics of the proteins extracted from complex microbial consortia in biofilm with extraction buffers containing different detergents
Summary
Biofilms are the structured consortia of trophic groups of microorganisms grown in a close approximation on the surface of supporting media, which can be involved an important biological process for water/wastewater purification, bioenergy recovery, and environmental remediation. Similar to any other proteomic experiments, a metaproteomic analysis undergoes protein recovery from the studied materials, enzymatic digestion, mass-based protein identification, and annotation to generate a dataset that can provide intensive insight to microbial community composition, functionality and activity. The proteome from a biofilm sample is inherently complicated, because the proteins of different abundance are produced by numerous microorganisms with highly complex matrix of extracellular polymeric substances. In this regard, the sample preparation such as protein extraction and purification to achieve efficiency for the environmental samples is very challenging [7,10]. We systematically evaluated the effects of detergents of different types, such as anionic, neutral, zwitterionic and mixed, on recovering the proteins from biofilm of an anaerobic bioreactor
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