N-Acyl-N-alkyl/aryl Sulfonamide Chemistry Assisted by Proximity for Modification and Covalent Inhibition of Endogenous Proteins in Living Systems.

Abstract

ConspectusSelective chemical modification of endogenous proteins in living systems with synthetic small molecular probes is a central challenge in chemical biology. Such modification has a variety of applications important for biological and pharmaceutical research, including protein visualization, protein functionalization, proteome-wide profiling of enzyme activity, and irreversible inhibition of protein activity. Traditional chemistry for selective protein modification in cells largely relies on the high nucleophilicity of cysteine residues to ensure target-selectivity and site-specificity of modification. More recently, lysine residues, which are more abundant on protein surfaces, have attracted attention for the covalent modification of proteins. However, it has been difficult to efficiently modify the ε-amino groups of lysine side-chains, which are mostly (∼99.9%) protonated and thus exhibit low nucleophilicity at physiological pH. Our group revealed that N-acyl-N-alkyl sulfonamide (NASA) moieties can rapidly and efficiently acylate noncatalytic (i.e., less reactive) lysine residues in proteins by leveraging a reaction acceleration effect via proximity. The excellent reaction kinetics and selectivity for lysine of the NASA chemistry enable covalent modification of natural intracellular and cell-surface proteins, which is intractable using conventional chemistries. Moreover, recently developed N-acyl-N-aryl sulfonamide (ArNASA) scaffolds overcome some problems faced by the first-generation NASA compounds. In this Account, we summarize our recent works in the development of NASA/ArNASA chemistry and several applications reported by ourselves and other groups. First, we characterize the basic properties of NASA/ArNASA chemistry, including the labeling kinetics, amino acid preference, and biocompatibility, and compare this approach with other ligand-directed chemistries. This section also describes the principles of nucleophilic organocatalyst-mediated protein acylation, another important protein labeling strategy using the NASA reactive group, and its application to neurotransmitter receptor labeling in brain slices. Second, we highlight various recent examples of protein functionalization using NASA/ArNASA chemistry, such as visualization of membrane proteins including therapeutically important G-protein coupled receptors, gel-based ligand screening assays, photochemical control of protein activity, and targeted protein degradation. Third, we survey covalent inhibition of proteins by NASA/ArNASA-based lysine-targeting. The unprecedented reactivity of NASA/ArNASA toward lysine allows highly potent, irreversible inhibition of several drug targets for the treatment of cancer, including HSP90, HDM2-p53 protein-protein interaction, and a Bruton's tyrosine kinase mutant that has developed resistance to cysteine-targeted covalent-binding drugs. Finally, current limitations of, and future perspectives on, this research field are discussed. The new chemical labeling techniques offered by NASA/ArNASA chemistry and its derivatives create a valuable molecular toolbox for studying numerous biomolecules in living cells and even in vivo.