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Covalent chemical probes – chemical probes that form a covalent bond with their target biomacromolecule(s) represent powerful tools that can be used for biology discovery, target validation (or off-target identification), and as starting points for drug discovery programmes.
Drugs with a covalent mode of action have been known and used for over a century, although historically, pursuit of such bioactive compounds has been avoided due to concerns over lack of selectivity. Over the last three decades, however, the use of ligands bearing reactive handles has seen increased interest spawning efforts to rationally design covalent drugs and new approaches to study protein function such as activity-based protein-profiling. Where covalent bioactive ligands are concerned, increased selectivity and duration of action represent advantages. In the context of new methods to study biology, chemical methods uniquely offer the ability to develop new reactive warheads, optimize bioorthogonal chemistries, and label a protein-of-interest with a (multi)functional group of choice, including natural or non-natural functionalities harbouring a recognition, reporting or affinity handle. Collectively, this offers new opportunities to modulate, track and isolate proteins of interest in/from the complex cellular milieu. In turn, this has created demand for new regioselective chemistries and analytical methods to assess biomacromolecule labelling in the complex environment of the cell.
This Guest Edited Collection aims to bring together research across the broad remit of covalent biomacromolecule labelling chemistry. We welcome both experimental and theoretical studies, with topics of interest including but not limited to:
New chemistry for protein modification
Photoaffinity labelling
Protein target identification
Chemical proteomics or chemoproteomics
Activity based protein profiling
Proximity-based labelling
Bioorthogonal chemistry
Covalent inhibitors
Covalent degraders
Understanding post-translational modifications using chemical approaches
In addition to primary research Articles, we also welcome Perspectives, Reviews, and Comments. All submissions will be subject to the same review process and editorial standards as regular Communications Chemistry Articles.
Developing effective inhibitors of the DNA repair enzyme tyrosyl-DNA phosphodiesterase 1 (TDP1) is challenging because of the enzyme’s shallow catalytic pocket and non-specific substrate binding interactions. Here, the authors use Sulfur (VI) fluoride exchange chemistries to prepare covalent TDP1-bound binders showing site-specific covalent bonds with the Y204 residue that position DNA.
Immune-cell reprogramming driven by mitochondria-derived reactive electrophilic immunometabolites (mt-REMs) is an emerging phenomenon of major biomedical importance. Here, the authors highlight the latest advances and overarching challenges in precision indexing of mt-REMs’ cellular responses with spatiotemporal intelligence and locale-specific function assignments.
Multifunctional cysteine targeting covalent warheads possess significant therapeutic potential in medicinal chemistry and chemical biology. Here, the authors develop an oxazolinosene scaffold from nitrile groups and saccharides that can selectively conjugate cysteine residues within peptides and proteins under physiological conditions, as well as deplete glutathione in cancer cells.
Site-specific conjugation of oligonucleotides and native proteins remains challenging. Here, the authors select covalent DNA aptamers from a library modified with N-hydroxysuccinimide esters, and show their application in the formation of antibody–oligonucleotide conjugates for protein detection.
Fragment screening is a popular strategy for generating viable chemical starting points for drug targets, however, weak affinity to targets, as well as the exploration of the binding site, remain challenging. Here, the authors develop pharmacophore-optimized photoaffinity fragments that can effectively explore the available binding site and enable a high hit rate and better sensitivity.
Histone methylation by histone lysine methyltransferases (HKMTs) is a vital post-translational modification regulating gene expression, however, selective mapping of methylation by proteomics analysis remains challenging. Here, the authors develop a heavy co-factor analogue 13CD3-BrSAM for HKMT DOT1L that can selectively heavy label target substrates, and map their methylation by proteomics.
The binding and phosphorylation of serine–arginine-rich (SR) proteins by SR protein kinases (SRPKs) is essential to regulate target gene expression, however, the efficient inhibition of this interaction and phosphorylation remains underexplored. Here, the authors develop a covalent inhibitor that targets the lysine residue within the SRPK-specific docking groove, to block interaction and phosphorylation of the prototypic SR protein SRSF1.
Mass spectrometry-based quantitative chemoproteomics is widely used for the identification of protein targets as well as modified residues, however, sample preparation and data analysis remain tedious. Here, the authors develop silane-based cleavable linkers functionalized tandem mass tags as click-compatible isobaric tags, introducing the isobaric label earlier in sample preparation, achieving decreased sample preparation time, with high coverage and high-accuracy quantification.
Covalent probes are a powerful tool for investigating small molecule and protein interactions, however, the development of reactive warheads to form covalent probes remains underexplored. Here, the authors develop α-acyloxyenamide electrophiles to covalently bind to lysine residues, and selectively profile the conserved lysine residues of kinase in live cells.
Protease inhibitors represent attractive antiviral agents against coronaviruses, however, current inhibitors show restricted efficiency which limits their use. Here, the authors develop covalent reversible peptidyl inhibitors with nitroalkene warheads, showing inhibition against main protease and human Cathepsin L, preventing viral infection in a cellular assay.
Fluorinated ketones can function as covalent warheads applied in the design of reversible covalent inhibitors, however, the reactivity of certain fluorinated moieties remains underexplored. Here, the authors investigate the species-specific lipophilicities of fluorinated geminal diketones in multicomponent equilibrium systems and reveal their potential as multifaceted warheads for reversible covalent drugs.
mRNA display strategies such as random non-standard peptide integrated discovery (RaPID) system allow screening of large peptide libraries to identify reversible binding to a target of interest. Here, the authors develop a covalent version of this methodology, photocrosslinking-RaPID, allowing identification of peptides that efficiently covalently modify their target of choice via photoaffinity labelling.
Excavatolide B (excB) is a marine briarane type diterpenoid with anti-inflammatory properties, however, its cellular targets and mode-of-action remain unknown. Here, the authors develop two covalent probes of excB and apply them through a chemoproteomics approach to identify STING as a direct target of excB in living mammalian cells.
Fibroblast growth factor receptor 4 (FGFR4) is a promising target for the treatment of hepatocellular carcinoma, but current FGFR4 covalent inhibitors target only one of the two cysteine residues (Cys477 or Cys552) that provide FGFR4-specificity. Here, a dual-warhead covalent FGFR4 inhibitor that can covalently target both cysteine residues of FGFR4 is reported, and strong selectivity for FGFR4 is observed.
Proximity-based ligations commonly require an external stimulus such as a catalyst or irradiation, or highly reactive functional groups. Here the reaction of alpha effect nucleophiles and 2,5-dioxopentanyl derivatives allows direct proximity-based ligation while avoiding highly reactive moieties.
Strained alkenes are valuable reagents for rapid and selective labeling of biomolecules but may undergo side-reactions. Here direct excitation of an azobenzene generates a strained nitrogen-nitrogen double bond in situ which reacts with a photochemically-generated nitrile imine, allowing the labeling of live cells with spatiotemporal control.
Ligand-directed protein labeling allows selective modification of native proteins but typically requires stoichiometric quantities of the labeling agent. Here a substoichiometric quantity of a peptide probe bound to a photocatalyst allows selective labeling of a target protein Cys residue in the presence of structurally similar proteins.
Tumor cells express high levels of integrins, which can be labeled using cyclic peptides, but selective labeling of target cells can be a challenge. Here in situ ligation of strongly binding cyclic peptides with weakly binding glycans allows discrimination of cancerous and non-cancerous cells which both express integrins on the surface.
Oligomerisation is widely used to engineer proteins and peptides with desirable properties. Here, covalent homodimers and heterodimers of fluorescent proteins are designed in silico and experimentally shown to exhibit differing spectral properties depending upon the structure of the protein-protein interface.
Linear ubiquitylation of NEMO is implicated in intracellular immune signalling but monoubiquitylated NEMO is synthetically inaccessible. Here the authors report auxiliary-assisted ubiquitylation of peptides including the CoZi domain of NEMO and explore its linear ubiquitylation mechanism.