Diaryliodonium-based probes for activity-based protein profiling of oxidoreductases Leo Krammer 1 , Barbara Darnhofer 2 , Ruth Birner-Grünberger 2,3 , Rolf Breinbauer 1 1 Institute of Organic Chemistry, Graz University of Technology, Austria 2 Diagnostic and Research Institute of Pathology, Medical University of Graz, Austria 3 Institute of Chemical Technologies and Analytics, University of Technology Vienna, Austria Activity-based protein profiling (ABPP) represents an intriguing and powerful proteomic method for the in vitro and invivo identification and evaluation of proteins in its active state. [1] Throughout all enzyme classes, hydrolases can be considered as the so far most addressed enzymes, since they usually feature a nucleophilic residue (e.g. serine, cysteine), which can attack the electrophilic warhead of the activity-based probe (ABP). Oxidoreductases (EC 1), in contrast, have received much less attention, although they play a role in essential metabolic processes and a more detailed insight into their function and activity could be beneficial to the understanding of cellular biochemistry and the role of these enzymes in diseases. [2] Previously reported ABPs used for oxidoreductases include, amongst others, examples for the highly specific targeting of MAO [3,4] or more general approaches, for example towards cytochrome P450s. [5] However, it could also be desirable to label different subclasses of oxidoreductases in a single ABPP experiment with the same activity-based probe. In order to take on this challenge, we designed and synthesized several novel ABPs based on the potent ALDH2 inhibitor and hypervalent iodine compound diphenyleneiodonium (DPI) [6] and its analogue diphenyliodonium (IDP) (see Figure 1).
Figure 1. Schematic workflow for the activity-based protein profiling of oxidoreductases with diaryliodonium-based probes. The probes were then used for in vitro ABPP labeling experiments with fresh mouse liver. After proteome labeling and initial SDS-PAGE in-gel fluorescence assays, promising candidates were further used in biotin/streptavidin affinity enrichment, whereupon labeled proteins could be identified by LC-MS/MS after tryptic digestion. References 1. B. F. Cravatt, A. T. Wright, J. W. Kozarich, Annu. Rev. Biochem . 2008 , 77 , 383–414. 2. R. Fuerst, R. Breinbauer, ChemBioChem 2021 , 22 , 630–638. 3. J. M. Krysiak, J. Kreuzer, P. Macheroux, A. Hermetter, S. A. Sieber, R. Breinbauer, Angew. Chem. Int. Ed. 2012 , 51 , 7035–7040. 4. L. Li, C.-W. Zhang, J. Ge, L. Qian, B.-H. Chai, Q. Zhu, J.-S. Lee, K.-L. Lim, S. Q. Yao, Angew. Chem. Int. Ed. 2015 , 54 , 10821–10825. 5. A. T. Wright, B. F. Cravatt, Chem. Biol. 2007 , 14 , 1043–1051. 6. R. Neubauer, A. Neubauer, G. Wölkart, C. Schwarzenegger, B. Lang, K. Schmidt, M. Russwurm, D. Koesling, A. C. F. Gorren, A. Schrammel, B. Mayer, Mol. Pharmacol. 2013 , 84 , 407-414.
P45F
© The Author(s), 2023
Made with FlippingBook Learn more on our blog