Dataset Information

Total protein extraction for metaproteomics analysis of methane producing biofilm: the effects of detergents.

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.

SUBMITTER: Huang HJ

PROVIDER: S-EPMC4100146 | biostudies-literature | 2014 Jun

REPOSITORIES: biostudies-literature

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Total protein extraction for metaproteomics analysis of methane producing biofilm: the effects of detergents.

Huang Hung-Jen HJ Chen Wei-Yu WY Wu Jer-Horng JH

International journal of molecular sciences 20140606 6

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 ...[more]

PMID: 24914765

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Project description:Methane emissions from aquatic ecosystems are increasingly recognized as substantial, yet variable, contributions to global greenhouse gas emissions. This is in part due to the challenge of modeling biologic parameters that affect methane emissions from a wide range of sediments. For example, the impacts of fish bioturbation on methane emissions in the literature have been shown to result in a gradient of reduced to enhanced emissions from sediments. However, it is likely that variation in experimental fish density, and consequently the frequency of bioturbation by fish, impacts this outcome. To explore how the frequency of disturbance impacts the levels of methane emissions in our previous work we quantified greenhouse gas emissions in sediment microcosms treated with various frequencies of mechanical disturbance, analogous to different levels of activity in benthic feeding fish. Greenhouse gas emissions were largely driven by methane ebullition and were highest for the intermediate disturbance frequency (disturbance every 7 days). The lowest emissions were for the highest frequency treatment (3 days). This work investigated the corresponding impacts of disturbance treatments on the microbial communities associated with producing methane. In terms of total microbial community structure, no statistical difference was observed in the total community structure of any disturbance treatment (0, 3, 7, and 14 days) or sediment depth (1 and 3 cm) measured. Looking specifically at methanogenic Archaea however, a shift toward greater relative abundance of a putatively oxygen-tolerant methanogenic phylotype (ca. Methanothrix paradoxum) was observed for the highest frequency treatments and at depths impacted by disturbance (1 cm). Notably, quantitative analysis of ca. Methanothrix paradoxum demonstrated no change in abundance, suggesting disturbance negatively and preferentially impacted other methanogen populations, likely through oxygen exposure. This was further supported by a linear decrease in quantitative abundance of methanogens (assessed by qPCR of the mcrA gene), with increased disturbance frequency in bioturbated sediments (1 cm) as opposed to those below the zone of bioturbation (3 cm). However, total methane emissions were not simply a function of methanogen populations and were likely impacted by the residence time of methane in the lower frequency disturbance treatments. Low frequency mechanical disruption results in lower methane ebullition compared to higher frequency treatments, which in turn resulted in reduced overall methane release, likely through enhanced methanotrophic activities, though this could not be identified in this work. Overall, this work contributes to understanding how animal behavior may impact variation in greenhouse gas emissions and provides insight into how frequency of disturbance may impact emissions.

Dataset Information

Total protein extraction for metaproteomics analysis of methane producing biofilm: the effects of detergents.

Publications

Total protein extraction for metaproteomics analysis of methane producing biofilm: the effects of detergents.

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