The microbial content associated with humans is immense, amassing to almost equal amounts of microbial and human cells in our bodies. We are beginning to appreciate associations of certain microbes and microbial genes with particular host physiological states. However, considerable information is lacking about relevant functional activities of the human microbiome as the origin of certain metabolites and function of diverse enzymes is unknown. For example, more than 50% of genes in human gut metagenomes cannot be annotated and ca. 10% of mammalian plasma metabolites are thought to have a gut microbial component, which can be identified in less than 5% of these molecules. Such knowledge gaps hinder progress towards preventing and treating diseases linked to the gut microbiome, such as colorectal cancer. As a result, efforts to extend our understanding of the organisms present in these communities has increased.<br/>Glycyl radical enzymes (GREs), for example, are one of the more abundant protein families in the human gut microbiome, supporting metabolic steps in strict and facultative anaerobes. Harboring a stable free radical in their active state, GREs are important biocatalysts that generally occur as homodimeric enzymes with a subunit size of 80–100 kDa; and while numerous novel GREs are getting discovered, including benzylsuccinate synthase (BSS; EC 4.1.99.11) or 4-hydroxyphenylacetate decarboxylase (HPD; EC 4.1.1.82), the functions of other putative GREs remain unknown.<br/>In taking advantage of the unique reactivity of protein backbone-based radical intermediates, activity-based protein profiling probes are being developed to identify and characterize novel GREs, providing additional information on their functional states within intact microbial communities. To illustrate this strategy, we developed small molecule model reactions containing a single amino acid or a series of peptides homologous mimicking the active sites of selected GREs, in which we demonstrate that glycine-based carbon-centered radicals can be trapped and isolated in the presence of various functional groups.

Chemical tools to decipher gut microbial communities: Detection and characterisation of putative glycyl radical enzymes (GREs)

256th National Meeting (2018)

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