Valegård Karin, Terwisscha van Scheltinga Anke C, Dubus Alain, Ranghino Graziella, Oster Linda M, Hajdu Janos, Andersson Inger
Molecular Biophysics, Department of Cellular and Molecular Biology, Uppsala University, Box 596, S-751 24 Uppsala, Sweden.
Nat Struct Mol Biol. 2004 Jan;11(1):95-101. doi: 10.1038/nsmb712. Epub 2003 Dec 29.
Deacetoxycephalosporin-C synthase (DAOCS) is a mononuclear ferrous enzyme that transforms penicillins into cephalosporins by inserting a carbon atom into the penicillin nucleus. In the first half-reaction, dioxygen and 2-oxoglutarate produce a reactive iron-oxygen species, succinate and CO2. The oxidizing iron species subsequently reacts with penicillin to give cephalosporin and water. Here we describe high-resolution structures for ferrous DAOCS in complex with penicillins, the cephalosporin product, the cosubstrate and the coproduct. Steady-state kinetic data, quantum-chemical calculations and the new structures indicate a reaction sequence in which a 'booby-trapped' oxidizing species is formed. This species is stabilized by the negative charge of succinate on the iron. The binding sites of succinate and penicillin overlap, and when penicillin replaces succinate, it removes the stabilizing charge, eliciting oxidative attack on itself. Requisite groups of penicillin are within 1 A of the expected position of a ferryl oxygen in the enzyme-penicillin complex.
去乙酰氧基头孢菌素 - C 合酶（DAOCS）是一种单核亚铁酶，它通过将一个碳原子插入青霉素核中将青霉素转化为头孢菌素。在第一个半反应中，氧气和 2 - 氧代戊二酸产生一种活性铁 - 氧物种、琥珀酸和二氧化碳。随后，氧化性铁物种与青霉素反应生成头孢菌素和水。在此，我们描述了亚铁 DAOCS 与青霉素、头孢菌素产物、共底物和副产物形成复合物的高分辨率结构。稳态动力学数据、量子化学计算和新结构表明了一个反应序列，其中形成了一种“陷阱式”氧化性物种。该物种通过琥珀酸在铁上的负电荷得以稳定。琥珀酸和青霉素的结合位点重叠，当青霉素取代琥珀酸时，它消除了稳定电荷，引发对自身的氧化攻击。青霉素的必需基团位于酶 - 青霉素复合物中一个铁氧中间体预期位置的 1 埃范围内。
