Pyrroloquinoline quinone (PQQ) is a bacterial redox cofactor enabling enzyme catalysis in various sugar and alcohol dehydrogenases. However, its proposed additional role as a "longevity vitamin" lacks a clear molecular basis and is thus highly debated. Here, we applied chemical proteomics to identify previously unknown classes of PQQ-binding proteins. We designed and synthesized a structurally diverse suite of five PQQ probes equipped with a diazirine photo-cross-linker and an alkyne handle for target identification. The fidelity of the probes was first evaluated for two well-characterized bacterial PQQ-dependent enzymes, demonstrating not only probe binding but also the reconstitution of catalytic activity. We then commenced with proteome profiling of Escherichia coli and Pseudomonas putida cells and unraveled a distinct set of putative PQQ-binding proteins. Recombinant expression of selected hits, including several chaperones, validated PQQ binding. Notably, in some cases, PQQ even formed covalent adducts with selected lysine residues, for instance, in the AAA+ ATPase RuvB involved in DNA remodeling. Overall, our work highlights the utility of PQQ probes to further unravel the complement of cofactor-binding proteins in whole cells. It also provides a basis for future mechanistic studies of PQQ functions beyond redox catalysis.
Wang et al. (Thu,) studied this question.