Deconvoluting the biology and druggability of protein lipidation using chemical proteomics

Lipids are indispensable cellular building blocks, and their post-translational attachment to proteins makes them important regulators of many biological processes. Dysfunction of protein lipidation is also implicated in many pathological states, yet its systematic analysis presents significant challenges. Thanks to innovations in chemical proteomics, lipidation can now be readily studied by metabolic tagging using functionalized lipid analogs, enabling global profiling of lipidated substrates using mass spectrometry. This has spearheaded the first deconvolution of their full scope in a range of contexts, from cells to pathogens and multicellular organisms. Protein N-myristoylation, S-acylation, and S-prenylation are the most well-studied lipid post-translational modifications because of their extensive contribution to the regulation of diverse cellular processes. In this review, we focus on recent advances in the study of these post-translational modifications, with an emphasis on how novel mass spectrometry methods have elucidated their roles in fundamental biological processes

Characterisation of the key determinants of Phd antitoxin mediated Doc toxin inactivation in Salmonella

In the search for novel antimicrobial therapeutics, toxin-antitoxin (TA) modules are promising yet underexplored targets for overcoming antibiotic failure. The bacterial toxin Doc has been associated with the persistence of Salmonella in macrophages, enabling its survival upon antibiotic exposure. After developing a novel method to produce the recombinant toxin, we have used antitoxin-mimicking peptides to thoroughly investigate the mechanism by which its cognate antitoxin Phd neutralizes the activity of Doc. We reveal insights into the molecular detail of the Phd–Doc relationship and discriminate antitoxin residues that stabilize the TA complex from those essential for inhibiting the activity of the toxin. Coexpression of Doc and antitoxin peptides in Salmonella was able to counteract the activity of the toxin, confirming our in vitro results with equivalent sequences. Our findings provide key principles for the development of chemical tools to study and therapeutically interrogate this important class of protein–protein interactions