Design and engineering of a man-made diffusive electron-transport protein.
Design and engineering of a man-made diffusive electron-transport protein.
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DOI：
10.1016/j.bbabio.2015.09.008
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发表时间：
2016-05
期刊：
影响因子：
--
通讯作者：
Moser CC
中科院分区：
文献类型：
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作者：
Fry BA;Solomon LA;Leslie Dutton P;Moser CC
Maquettes 是人造辅因子结合氧化还原酶，根据基本原理设计，对天然蛋白质序列的参考极少。在这里，我们重点关注水溶性模型，其设计和工程用于执行通常由细胞色素、铁氧还蛋白和黄素氧还蛋白以及其他小蛋白质在光合作用和呼吸能量转换以及氧化还原代谢中进行的扩散电子传输。我们的设计通过分析血红素模型和众所周知的天然电子转运蛋白细胞色素 c 之间的电子转移进行了测试。通过停流测量从几秒到几毫秒的电子转移动力学，同时通过一氧化碳模型血红素复合物的激光光解实现亚毫秒分辨率。这些测量结果证明了电子从模型到细胞色素 c 的转移，再现了在自然系统中观察到的时间尺度和电荷互补性调制。蛋白质间电子转移从 9.7 × 106 M−1s−1 到 1.2 × 109 M−1s−1 的离子强度依赖性遵循简单的 Debye-Hückel 模型，用于 +8 净电荷氧化细胞色素 c 和 -19 净电荷血红素模型之间的吸引力，没有迹象表明存在显着的蛋白质偶极矩转向。成功地在人造蛋白质中重建能量转换和下游代谢的重要组成部分，为体内临床干预以及燃料或其他工业产品的生产带来了希望。
Maquettes are man-made cofactor-binding oxidoreductases designed from first principles with minimal reference to natural protein sequences. Here we focus on water-soluble maquettes designed and engineered to perform diffusive electron transport of the kind typically carried out by cytochromes, ferredoxins and flavodoxins and other small proteins in photosynthetic and respiratory energy conversion and oxido-reductive metabolism. Our designs were tested by analysis of electron transfer between heme maquettes and the well-known natural electron transporter, cytochrome c. Electron-transfer kinetics were measured from seconds to milliseconds by stopped-flow, while sub-millisecond resolution was achieved through laser photolysis of the carbon monoxide maquette heme complex. These measurements demonstrate electron transfer from the maquette to cytochrome c, reproducing the timescales and charge complementarity modulation observed in natural systems. The ionic strength dependence of inter-protein electron transfer from 9.7 × 106 M−1s−1 to 1.2 × 109 M−1s−1 follows a simple Debye-Hückel model for attraction between +8 net charged oxidized cytochrome c and −19 net charged heme maquette, with no indication of significant protein dipole moment steering. Successfully recreating essential components of energy conversion and downstream metabolism in man-made proteins holds promise for in vivo clinical intervention and for the production of fuel or other industrial products.