Department of Human and Animal Biology, University of Turin, Italy.
J Am Chem Soc. 2010 Jan 20;132(2):458-9. doi: 10.1021/ja909261p.
This communication reports on the first electrochemical study of the human flavin-containing monooxygenase 3 (hFMO3) either absorbed or covalently linked to different electrode surfaces. Glassy carbon and gold electrodes gave reversible electrochemical signals of an active hFMO3. The midpoint potential measured for the immobilized enzyme on a glassy carbon electrode was -445 +/- 8 mV (versus Ag/AgCl). A monolayer coverage was obtained on gold functionalized with dithio-bismaleimidoethane that covalently linked surface accessible cysteines of hFMO3. A structural model of the enzyme was generated to rationalize electrochemistry results. The turnover of the active enzyme was measured with two specific drugs: tamoxifen and benzydamine. For tamoxifen, 1.7 and 8.0 microM of its N-oxide product were formed by the enzyme immobilized on glassy carbon and gold electrodes, respectively. In the case of benzydamine, a K(M) of 44 +/- 5 microM was measured upon application of a -600 mV bias to the enzyme immobilized on the glassy carbon electrode that is in good agreement with the values published for microsomal hFMO3 where NADPH is the electron donor.
本通讯报道了人黄素单加氧酶 3（hFMO3）在被吸收或共价连接到不同电极表面时的首次电化学研究。玻碳和金电极给出了活性 hFMO3 的可逆电化学信号。在玻碳电极上固定化酶的中点电位测量值为-445 +/- 8 mV（相对于 Ag/AgCl）。用二硫代双马来酰亚胺乙烷功能化的金获得了单层覆盖，该化合物共价连接了 hFMO3 表面可及的半胱氨酸。生成了酶的结构模型以合理化电化学结果。使用两种特定药物测量了活性酶的周转率：他莫昔芬和苯佐卡因。对于他莫昔芬，在玻碳和金电极上固定化的酶分别形成了 1.7 和 8.0 microM 的其 N-氧化物产物。对于苯佐卡因，当在玻碳电极上固定化的酶上施加-600 mV 的偏压时，测量到的 K（M）为 44 +/- 5 microM，这与作为电子供体的 NADPH 的微粒体 hFMO3 发表的值非常吻合。
