A coumarin-porphyrin break-apart probe for heme oxygenase-1 Edward R. H. Walter 1,2 , Ying GE 2 , Saul M. Cooper 1,2 , Justin C. Mason 2 , Joseph J. Boyle 2 , Nicholas J. Long 1 1 Imperial College London, UK, 2 National Heart and Lung Institute, Imperial College London, UK Heme oxygenase-1 (HO-1) is an important enzyme in vascular biology that is primarily responsible for the regulation of cytotoxic free heme. 1 Heme catabolism is a regiospecific three-step process that requires molecular oxygen, NADPH and cytochrome p450 reductase, to form biliverdin and carbon monoxide. 2 HO-1 overexpression is commonly associated with a number of cardiovascular and neurogenerative diseases, and, therefore, has the potential to act as a diagnostic / prognostic marker in this regard. However, to-date HO-1 has been somewhat overlooked as a potential marker for vascular disease. We shall describe our recent work toward the design, synthesis, photophysical and biological characterisation of the first known chemical probe to report on HO-1 activity. 3,4 Probe Fe–L 1 was designed to utilise the regiospecific HO-1- catalysed porphyrin degradation of heme and ‘break-apart’, perturbing the efficient Fluorescence Resonance Energy Transfer (FRET) mechanism from a coumarin donor to a porphyrin accepter fluorophore ( Figure 1A ). Analysis of HO-1 activity in Escherichia coli lysates overexpressing HO-1, determined that porphyrin degradation was regiospecific at the α-position following incubation with NADPH, resulting in the formation of coumarin 1 and a 6-fold increase in fluorescence ( Figure 1C ). Through the analysis of Fe–L 2 ( Figure 1B ), it was confirmed that close structural analogues of heme are required to maintain HO‑1 activity. 3,4 It is anticipated that this proof-of-concept study will act as a foundation to develop new red-shifted probes to report on HO-1 activity in the near future.
Times New Roman', serif; color:#000000; font-variant-caps:normal; letter-spacing:normal; text-transform:none; word- spacing:0px; text-decoration:none; text-indent:35.4pt;" >
Figure 1. (A) HO-1 catabolism of Fe–L 1 and (B) control porphyrin Fe–L 2 . (C) The change in coumarin fluorescence at 383 nm of Fe–L 1 and Fe–L 2 in E. coli lysates with and without incubation with NADPH. 3,4 References 1. Yuan, X.; Rietzschel, N.; Kwon, H.; Nuno, A. B. W.; Hanna, D. A.; Phillips, J. D.; Raven, E. L.; Reddi, A. R.; Hamza, I. Proc. Natl. Acad. Sci. U. S. A , 2016 , 113, 5144 2. Ortiz De Montellano, P. R. Curr. Opin. Chem. Biol, 2000 , 4, 221. 3. Walter, E. R. H.; Ge, Y.; Mason, J. C.; Boyle, J. J.; Long, N. J. J. Am. Chem. Soc , 2021 , 143, 6460. 4. Boyle, J. J.; Long, N. L.; Walter, E. R. H.; Ge, Y.; Mason, J. C.; WO2022/101635 A1.
P20
© The Author(s), 2022
Made with FlippingBook Learn more on our blog