Small Molecule Modulation of the Atypical Kinase/ATPase COQ8A

Abstract

Small molecule manipulation of the protein kinase‐like (PKL) superfamily has emerged as a remarkably effective therapeutic strategy, with more than 70 new drugs approved in the past 20 years, and has aided mechanistic studies of PKL function. However, such tools are still lacking for many PKL members, including the growing number of atypical kinases and pseudokinases. Among these are the ABC1/ADCK proteins, which are conserved throughout all domains of life and are directly tied to human diseases. ADCKs have been implicated in various aspects of prenyl‐lipid biology, with the most established being a role for COQ8 in coenzyme Q (CoQ) biosynthesis. While the importance of these proteins is clear, they are largely understudied and a mechanistic understanding of their function is lacking. To address this, we developed a series of small molecule probes to manipulate COQ8 activity in vitroand in cells. Here, we first present a biophysical investigation into the activation of COQ8A by the CoQ precursor mimetic 2‐propylphenol (2‐PP) using 1H‐13C HMQC NMR and hydrogen‐deuterium exchange mass spectrometry. We found that 2‐PP modulates the highly conserved KxGQ domain to increase COQ8A nucleotide affinity and ATPase activity. Second, we present the development of a custom small molecule inhibitor of COQ8A. Guided by crystallography, activity assays, and cellular CoQ measurements, we repurposed a 4‐anilinoquinoline scaffold to selectively inhibit COQ8A within mitochondria. Our newfound chemical tools promise to lend new mechanistic insights into the activities of these widespread and understudied proteins, and to offer potential therapeutic strategies for human diseases connected to their dysfunction.