Activity-Based Protein Profiling—Enabling Multimodal Functional Studies of Microbial Communities

Whidbey, Christopher; Wright, Aaron

doi:10.1007/82_2018_128

Cited by 22 publications

(22 citation statements)

References 81 publications

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“…However to date, they have not been applied for the labelling of whole cells within complex microbial communities, mainly due to the presence of the microorganisms within a complex undefined matrix. This poses major challenges for employing the CuAAC reaction, probe penetration into the cells, and labelling specificity and efficiency (104). The ABPP-based protocol developed in this study, to our knowledge for the first time, overcomes these challenges and allows the fluorescent visualization of catalytically active enzymes in situ .…”

Section: Discussionmentioning

confidence: 99%

An activity-based labelling method for the detection of ammonia and methane-oxidizing bacteria

Sakoula

Smith

Frank

et al. 2021

Preprint

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The advance of metagenomics in combination with intricate cultivation approaches has facilitated the discovery of novel ammonia- and methane-oxidizing microorganisms, indicating that our understanding of the microbial biodiversity within the biogeochemical nitrogen and carbon cycles still is incomplete. However, the in situ detection and phylogenetic identification of novel ammonia- and methane-oxidizing bacteria remains a challenge. Here, we describe an activity-based protein profiling protocol allowing cultivation-independent unveiling of ammonia- and methane-oxidizing bacteria. In this protocol, 1,7-octadiyne is used as a bifunctional enzyme probe that, in combination with a highly specific alkyne-azide cycloaddition reaction, enables the fluorescent or biotin labelling of cells harboring active ammonia and methane monooxygenases. The biotinylation of these enzymes in combination with immunogold labelling reveals the subcellular localization of the tagged proteins, while the fluorescent labelling of cells harboring active ammonia or methane monooxygenases provides a direct link of these functional lifestyles to phylogenetic identification when combined with fluorescence in situ hybridization. Furthermore, we show that this activity-based labelling protocol can be successfully coupled with fluorescence-activated cell sorting for the enrichment of nitrifiers and methanotrophs from complex environmental samples, facilitating the retrieval of their high quality metagenome-assembled genomes. In conclusion, this study demonstrates a novel, functional tagging technique for the reliable detection, identification, and enrichment of ammonia- and methane-oxidizing bacteria present in complex microbial communities.

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Section: Discussionmentioning

confidence: 99%

An activity-based labelling method for the detection of ammonia and methane-oxidizing bacteria

Sakoula

Smith

Frank

et al. 2021

Preprint

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“…Multiple enzymes from a single complex sample can, therefore, be identified based on their function ( Figure 4 A). Thus, activity-based protein profiling (ABPP) can enable functional discovery in complex, poorly annotated biological systems, including plants and soil microbial communities [ 1 , 36 , 37 ]. Even in one of the most studied model plants, Arabidopsis thaliana , only ~77% of the protein-coding sequences have been assigned structured annotation [ 38 ], while functional annotation of soil microbial communities remains a major experimental and computational challenge [ 39 , 40 , 41 ].…”

Section: Chemical Biology Toolboxmentioning

confidence: 99%

Interrogating Plant-Microbe Interactions with Chemical Tools: Click Chemistry Reagents for Metabolic Labeling and Activity-Based Probes

Lin

2021

Molecules

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Continued expansion of the chemical biology toolbox presents many new and diverse opportunities to interrogate the fundamental molecular mechanisms driving complex plant–microbe interactions. This review will examine metabolic labeling with click chemistry reagents and activity-based probes for investigating the impacts of plant-associated microbes on plant growth, metabolism, and immune responses. While the majority of the studies reviewed here used chemical biology approaches to examine the effects of pathogens on plants, chemical biology will also be invaluable in future efforts to investigate mutualistic associations between beneficial microbes and their plant hosts.

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“…The need to provide direct attribution of microbiota-derived metabolites to specific taxa has brought activity-based protein profiling (ABPP) to the forefront of microbiome science (Whidbey and Wright, 2019 ; Keller et al, 2020 ). ABPP exclusively selects for active proteins through function-dependent covalent labeling with small molecule activity-based probes (ABPs), (Cravatt et al, 2008 ).…”

Section: Functional Characterization Of the Gut Microbiome Using Actimentioning

confidence: 99%

Who Is Metabolizing What? Discovering Novel Biomolecules in the Microbiome and the Organisms Who Make Them

Couvillion

Agrawal

Colby

et al. 2020

Front. Cell. Infect. Microbiol.

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Even as the field of microbiome research has made huge strides in mapping microbial community composition in a variety of environments and organisms, explaining the phenotypic influences on the host by microbial taxa—both known and unknown—and their specific functions still remain major challenges. A pressing need is the ability to assign specific functions in terms of enzymes and small molecules to specific taxa or groups of taxa in the community. This knowledge will be crucial for advancing personalized therapies based on the targeted modulation of microbes or metabolites that have predictable outcomes to benefit the human host. This perspective article advocates for the combined use of standards-free metabolomics and activity-based protein profiling strategies to address this gap in functional knowledge in microbiome research via the identification of novel biomolecules and the attribution of their production to specific microbial taxa.

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Activity-Based Protein Profiling—Enabling Multimodal Functional Studies of Microbial Communities

Cited by 22 publications

References 81 publications

An activity-based labelling method for the detection of ammonia and methane-oxidizing bacteria

An activity-based labelling method for the detection of ammonia and methane-oxidizing bacteria

Interrogating Plant-Microbe Interactions with Chemical Tools: Click Chemistry Reagents for Metabolic Labeling and Activity-Based Probes

Who Is Metabolizing What? Discovering Novel Biomolecules in the Microbiome and the Organisms Who Make Them

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