The critical iron-oxygen intermediate in human aromatase |
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Authors: | Stephanie L Gantt Yelena V Grinkova |
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Institution: | a Department of Biochemistry, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA b Department of Chemistry, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA |
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Abstract: | Aromatase (CYP19) is the target of several therapeutics used for breast cancer treatment and catalyzes the three-step conversion of androgens to estrogens, with an unusual C-C cleavage reaction in the third step. To better understand the CYP19 reaction, the oxy-ferrous complex of CYP19 with androstenedione substrate was cryotrapped, characterized by UV-vis spectroscopy, and cryoreduced to generate the next reaction cycle intermediate. EPR analysis revealed that the initial intermediate observed following cryoreduction is the unprotonated g1 = 2.254 peroxo-ferric intermediate, which is stable up to 180 K. Upon gradual cryoannealing, the low-spin (g1 = 2.39) product complex is formed, with no evidence for accumulation of the g1 = 2.30 hydroperoxo-ferric intermediate. The relative stabilization of the peroxo-ferric heme and the lack of observed hydroperoxo-ferric heme distinguish CYP19 from other P450s, suggesting that the proton delivery pathway is more hindered in CYP19 than in most other P450s. |
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Keywords: | Cytochrome P450 Aromatase Peroxo-ferric intermediate Steroid biosynthesis EPR (electronic paramagnetic resonance) Oxy-ferrous heme complex |
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