Activity-based protein profiling in microbes and the gut microbiome

Abstract

Activity-based protein profiling (ABPP) is a powerful chemical approach for probing protein function and enzymatic activity in complex biological systems. This strategy typically utilizes activity-based probes that are designed to bind a specific protein, amino acid residue, or protein family and form a covalent bond through a reactivity-based warhead. Subsequent analysis by mass spectrometry-based proteomic platforms that involve either click chemistry or affinity-based labeling to enrich for the tagged proteins enables identification of protein function and enzymatic activity. ABPP has facilitated elucidation of biological processes in bacteria, discovery of new antibiotics, and characterization of host-microbe interactions within physiological contexts. This review will focus on recent advances and applications of ABPP in bacteria and complex microbial communities.

Keywords: Activity-based probes; Bacteria; Chemoproteomics; Click chemistry; Gut microbiome.

Figure 1.

Activity-based protein profiling (ABPP) can be used to identify enzymatic activities, protein function, and target proteins of small molecules using activity-based probes (ABPs). (a) Typical design of an ABP containing a reactive covalent warhead or electrophilic group, affinity element, and a click chemistry handle or reporter tag. (b) General workflow of ABPP or competitive ABPP using copper(I)-catalyzed azide-alkyne cycloaddition (CuAAC) to identify enzymatic activities and ABP target proteins using in-gel visualization and mass spectrometry-based proteomics.

Figure 2.

Chemical mechanisms of ABPs highlighted in this review. (a) Aldehydebased ABP for targeting protein lipoylation [19]. TCEP = tris(2-carboxyethyl)phosphine. (b) Light-induced ABP for capturing carboxylic acids in proteins [27]. (c) Mechanism based ABP targeting β-glucuronidases [–56].