Targeting biomolecules with reversible covalent chemistry

Abstract

Interaction of biomolecules typically proceeds in a highly selective and reversible manner, for which covalent bond formation has been largely avoided due to the potential difficulty of dissociation. However, employing reversible covalent warheads in drug design has given rise to covalent enzyme inhibitors that serve as powerful therapeutics, as well as molecular probes with exquisite target selectivity. This review article summarizes the recent advances in the development of reversible covalent chemistry for biological and medicinal applications. Specifically, we document the chemical strategies that allow for reversible modification of the three major classes of nucleophiles in biology: thiols, alcohols and amines. Emphasis is given to the chemical mechanisms that underlie the development of these reversible covalent reactions and their utilization in biology.

Figure 1

Comparison of a) noncovalent, b) irreversible covalent and c) reversible covalent inhibitors.

Figure 2

Reversible covalent chemistry of thiols. a) Reversible conjugation between an α-cyanoacrylate and cysteine. b) Crystal structure of a reversible covalent inhibitor bound to the kinase RSK2, highlighting the covalent linkage as well as a series of noncovalent interactions that give rise to target specificity. c) End-capped α-cyanoacrylamide inhibitors of the BTK kinase and crystal structure of 2 bound to BTK. d) Benzylidene rhodamine-based reversible covalent inhibitors of Hepatitis C NS5b RNA polymerase (left) and a cocrystal structure (right) to illustrate the covalent linkage. The structural images in b, c, and d are adapted with permission from references [7], [10] and [12] respectively.

Figure 3

Reversible covalent chemistry of alcohols. a) Reversible targeting of serine/threonine using boronic acid. b) Chemical structure of bortezomib, and its cocrystal structure with the yeast 20S proteasome (PDB: 2F16). c) Mechanism of action (left) and examples (right) of α-ketoamide-based reversible inhibitors. d) Three different classes of nitrile derivatives used for covalent drug design. e) Structure of saxagliptin as an example of nitrile-based reversible covalent inhibitors. f) Crystal structure of DPP4 showing covalent modification of Ser592 by saxagliptin.

Figure 4

Reversible covalent chemistry of amines. a) PLP-elicited imine formation. b) Coumarin 3-aldehyde based sensors for the detection of important amine-presenting metabolites. The table presents the substrate specificity of several sensors. c) A boronic acid-derivatized coumarin 3-aldehyde probe selectively binds glucosamine through cooperative imine and boronate ester formation. d) A molecular probe for the detection of histone deacetylase activity in live cells. e) Reversible iminoboronate formation of biological amines. f) Cartoon illustration of lysine side modification of proteins through iminoboronate formation. g) Illustration of bacterial labeling through iminoboronate formation of bacterial lipids. Shown on the right is a confocal image showing membrane labeling of S. aureus cells via iminoboronate chemistry.