Cellular production of a de novo membrane cytochrome.
Cellular production of a de novo membrane cytochrome.
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DOI：
10.1073/pnas.2300137120
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发表时间：
2023-04-18
影响因子：
11.1
通讯作者：
Curnow, Paul
中科院分区：
文献类型：
--
作者：
Hardy, Benjamin J.;Hermosilla, Alvaro Martin;Chinthapalli, Dinesh K.;V. Robinson, Carol;Anderson, J. L. Ross;Curnow, Paul
生命由光合作用和呼吸作用提供能量。这两个过程都依赖于完整膜蛋白的电子传递。这种生物能量蛋白的合成模型有助于揭示其折叠和组装背后的一般原理，并可用于构建体外和体内的替代电子传递链。在这里，我们使用计算设计来生产一种全新的膜细胞色素。该设计金属蛋白由重组细菌生物合成，自发获得氧化还原辅助因子血红素两分子，并能与其他蛋白质和小分子进行电子传递反应。因此，我们合理设计的蛋白质包含了生物能膜蛋白的几个核心特征，并在合成生物学中具有多种可能的应用。含血红素的整体膜蛋白是许多生物能量复合物和电子传递链的核心。这些电子中继中心在生物学中的重要性激发了重新设计蛋白质的灵感，这些蛋白质在健壮的、通用的和可处理的蛋白质折叠中重建了它们的核心特征。为此，我们在此报告了一种最小二血红素膜细胞色素的计算设计和细胞内生产，该细胞色素成功地整合到活细菌的细胞膜中。这种合成结构模拟了在现代呼吸复合物中发现的四螺旋束，但与自然界中发现的任何多肽序列没有序列同源性。这两种b型血红素似乎来自内源性血红素库，它们具有不同的分裂氧化还原电位，其值接近天然跨膜细胞色素的值。纯化后的蛋白质可以与小分子、其他氧化还原蛋白以及生物相关的扩散电子载体进行快速仿生电子传递。因此，我们报道了一种人工膜金属蛋白，它具有在合成原始细胞和生命系统中作为功能性电子传递模块的潜力。
Life is powered by photosynthesis and respiration. Both of these processes rely upon electron transport by integral membrane proteins. Synthetic models of such bioenergetic proteins help reveal the general principles behind their folding and assembly and could be used to build alternative electron transport chains in vitro and in vivo. Here, we use computational design to produce a de novo membrane cytochrome. This designer metalloprotein is biosynthesized by recombinant bacteria, spontaneously acquires two molecules of the redox cofactor heme, and can engage in electron transport reactions with other proteins and small molecules. Our rationally designed protein thus encapsulates several core features of bioenergetic membrane proteins and has multiple possible applications in synthetic biology. Heme-containing integral membrane proteins are at the heart of many bioenergetic complexes and electron transport chains. The importance of these electron relay hubs across biology has inspired the design of de novo proteins that recreate their core features within robust, versatile, and tractable protein folds. To this end, we report here the computational design and in-cell production of a minimal diheme membrane cytochrome which successfully integrates into the cellular membrane of live bacteria. This synthetic construct emulates a four-helix bundle found in modern respiratory complexes but has no sequence homology to any polypeptide sequence found in nature. The two b-type hemes, which appear to be recruited from the endogenous heme pool, have distinct split redox potentials with values close to those of natural membrane-spanning cytochromes. The purified protein can engage in rapid biomimetic electron transport with small molecules, with other redox proteins, and with biologically relevant diffusive electron carriers. We thus report an artificial membrane metalloprotein with the potential to serve as a functional electron transfer module in both synthetic protocells and living systems.
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影响因子：
15
作者：
Ghirlanda, G;Osyczka, A;DeGrado, WF
通讯作者：
DeGrado, WF
影响因子：
4.3
作者：
Alford RF;Koehler Leman J;Weitzner BD;Duran AM;Tilley DC;Elazar A;Gray JJ
通讯作者：
Gray JJ
影响因子：
14.8
作者：
Hernandez, Helena;Robinson, Carol V.
通讯作者：
Robinson, Carol V.
影响因子：
3.4
作者：
Alford, Rebecca F.;Fleming, Patrick J.;Gray, Jeffrey J.
通讯作者：
Gray, Jeffrey J.
影响因子：
2.9
作者：
Gibney, BR;Isogai, Y;Dutton, PL
通讯作者：
Dutton, PL