Genetically Controlled Syntheses of New Polymeric Materials

该项目将开发结构明确的人造蛋白质的制备方法，这些蛋白质携带活性烯烃、二烯基、乙炔和吡咯基官能团。该项目的一个关键方面是使用人工基因来指导均匀聚合物链的合成，这些聚合物链将直接（通过掺入非天然氨基酸）或间接（通过翻译后修饰反应）获得反应性功能。在这两种方法中，细菌蛋白质合成的保真度将被用于生产具有精确控制链长、序列和立体化学特征的聚合物材料。直接方法将探索与异亮氨酸、苯丙氨酸、组氨酸或酪氨酸相关的六种人工氨基酸（及其几种变体）的翻译活性。越来越多的证据表明，细菌蛋白质合成机制可以利用比通常由信使RNA模板编码的20种氨基酸更广泛的氨基酸范围。本研究选择的氨基酸类似物之所以特别有趣，是因为两个事实：1)天然氨基酸中没有烯基、二烯基、乙基或吡啶基；2)这些官能团可以用来赋予聚合物材料有趣的和潜在有用的化学和电化学性质。用于直接合并的方法包括：i)合成编码感兴趣序列的人工基因，用编码的天然类似物代替目标非天然氨基酸，ii)在适当的营养不良细菌菌株中克隆人工基因，以及iii)在含有非天然氨基酸（通常耗尽天然类似物）的培养基中诱导目标蛋白质合成。该策略已成功地用几种非天然氨基酸进行了验证，包括硒代蛋氨酸、对氟苯丙氨酸、3-噻吩丙氨酸、三氟亮氨酸、氮杂二羧酸、脱氢脯氨酸和硫脯氨酸。还将探讨涉及翻译后修饰的互补策略。在这种方法中，蛋白质生物合成将用于制备适当设计的“基础聚合物”，这些聚合物将用携带烯烃、二烯基、乙炔或吡咯基的试剂进行修饰。通过任何一种途径制备的聚合物都将受到分子结构、超分子组织和功能特性的仔细分析。特别关注的将是创造含有吡咯基侧链的电化学可切换蛋白质薄膜的前景。这种薄膜作为传感器、致动器和控制输送装置的元件具有潜在的实际用途。这项研究旨在通过使用人工基因来合成具有高度控制结构的氨基酸聚合物。所选择的聚合物在自然界中不存在，将被设计成具有有趣的电学和其他特性。* * *

9510031 Tirrell This project will develop methods for the preparation of architecturally well-defined artificial proteins carrying reactive olefinic, dienyl, acetylenic and pyrrolyl functional groups. A key aspect of the project is the use of artificial genes to direct the synthesis of uniform polymer chains, which will acquire reactive functionality either directly (through the incorporation of unnatural amino acids), or indirectly (through post-translational modification reactions). In either approach, the fidelity of bacterial protein synthesis will be exploited to produce polymeric materials characterized by precise control of chain length, sequence, and stereochemistry. The direct approach will explore the translational activity of six artificial amino acids (and several variants thereof) related to isoleucine, phenylalanine, histidine or tyrosine. There is increasing evidence that the bacterial protein synthesis machinery can utilize a braoder range of amino acids than the twenty that are normally encoded by messenger RNA templates. The amino acid analogues chosen for this study are of particular interest owing to two facts: i). none of the natural amino acids carries olefinic, dienyl, acetylenic or pyrrolyl groups, and ii). such functional groups can be used to endow polymeric materials with interesting and potentially useful chemical and electrochemical properties. The approach to be used for direct incorporation involves: i). synthesis of an artificial gene encoding the sequence of interest, with the natural analogue encoded in place of the target unnatural amino acid, ii). cloning of the artificial gene in an appropriate auxotrophic bacterial strain, and iii). induction of target protein synthesis in a medium containing the unnatural amino acid (and generally depleted of the natural analogue). This strategy has been demonstrated successfully with several unnatural amino acids, including selenomethionine, p-fluorophenylalanine, 3- thienylalanine, trifluoroleucine, azetidinecarboxylic acid, dehydroproline, and thiaproline. Complementary strategy involving post-translational modification will also be explored. In this approach, protein biosynthesis will be used to prepare appropriately designed "base polymers," which will be modified with reagents carrying olefinic, dienyl, acetylenic or pyrrolyl groups. Polymers prepared by either route will be subjected to careful analyses of molecular structure, supramolecular organization, and functional properties. Particular attention will be given to the prospects for creating electrochemically switchable protein films containing pyrrolyl side chains. Such films would be of potential practical use as components of sensors, actuators and controlled delivery devices. %%% This research aims to carry out the synthesis of amino acid polymers with highly controlled structures, through the use of artificial genes. The polymers selected do not occur in nature and will be designed to have interesting electrical and other properties. ***