Metaproteomic analysis of atmospheric aerosol samples

Abstract

Metaproteomic analysis of air particulate matter provides information about the abundance and properties of bioaerosols in the atmosphere and their influence on climate and public health. We developed and applied efficient methods for the extraction and analysis of proteins from glass fiber filter samples of total, coarse, and fine particulate matter. Size exclusion chromatography was applied to remove matrix components, and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) was applied for protein fractionation according to molecular size, followed by in-gel digestion and LC-MS/MS analysis of peptides using a hybrid Quadrupole-Orbitrap MS. Maxquant software and the Swiss-Prot database were used for protein identification. In samples collected at a suburban location in central Europe, we found proteins that originated mainly from plants, fungi, and bacteria, which constitute a major fraction of primary biological aerosol particles (PBAP) in the atmosphere. Allergenic proteins were found in coarse and fine particle samples, and indications for atmospheric degradation of proteins were observed.Graphical abstractWorkflow for the metaproteomic analysis of atmospheric aerosol samplesElectronic supplementary materialThe online version of this article (doi:10.1007/s00216-016-9747-x) contains supplementary material, which is available to authorized users.

Development of an Analytical Method for the Metaproteomic Investigation of Bioaerosol from Work Environments

“…The disruption of the cell wall is a critical phase of the extraction procedure for proteins collected on air filters, as PBAs are essentially made of fungal spores, bacteria, and other cells and cellular fragments. Compared to the previous paper already published on bioaerosol metaproteomics, where the authors presented an analytical method primarily aimed at the extraction of proteins that are easily released from PBAs, the extraction method tested in this work on B. subtilis did not only use a detergent and surfactant, but also included a mechanical lysis step for the disruption of microbial cell walls and spores present in BPAs . By this approach, the characteristic proteins of B. subtilis were identified by bioinformatics analysis, thus confirming the efficacy of the selected extraction method (data and chromatograms are reported in , Supporting Information).…”

“…Protocols A and C only differed for the use of EDTA to chelate metals, such as heavy metals, that could occur in air samples due to their ubiquitous and persistent nature. Protocol B was tested because it was the best protocol employed in a previous metaproteomic work; protocol B exploited a Tris/Gly/SDS buffer, which is suitable for the extraction of both water‐soluble and water‐insoluble proteins while minimizing other potential non‐covalent interactions, such as the ones between proteins and ambient aerosol components (e.g., soot, dust) or between proteins and the filter material. All three protocols required a protein precipitation step to remove detergents incompatible with the downstream protein quantification, digestion, and MS analysis.…”

“…In protocol A, the extraction buffer was prepared as previously described, with some modifications, and was made up of 15 mmol L −1 KCl, 2% w/v sodium dodecyl sulfate (SDS), 10 mmol L −1 ethylenediaminetetraacetic acid (EDTA), and 20 mmol L −1 dithiothreitol. Protocol B was based on an optimization of protein extraction from air filter samples developed in a previous work . The buffer used in protocol B contained 192 mmol L −1 glycine and 0.1% SDS in 25 mmol L −1 Tris‐HCl.…”

“…To date, most research examining protein PBAs are based on protein quantification by traditional assays, for example, bicinchoninic acid (BCA) assay, nano‐orange, and Bradford assay. The only metaproteomic paper already published was focused on ambient aerosol samples collected on the roof of the Max Planck Institute for Chemistry and exploited a gel‐based, time‐consuming proteomic approach which provided a very small number of protein identifications. Shotgun gel‐free separation is nowadays the best choice to analyze highly complex protein mixtures as it provides, compared to gel‐based approaches, a considerable reduction to the complexity of the sample without any loss of information .…”

Proteins and Amino Acids in Fine Particulate Matter in Rural Guangzhou, Southern China: Seasonal Cycles, Sources, and Atmospheric Processes

“…The discrepancy could be derived from the removal of LMW interfering substances before BCA analysis. It has been suggested that some LMW substances may react with BCA reagents, such as humic acid and glucose. , A size-exclusion column is suitable for removing interferences such as soot, ammonium sulfate, and humic-like substances (HULIS), and, most importantly, FAAs, , which were also analyzed in this study.…”