In Situ Deprotection and Incorporation of Unnatural Amino Acids during Cell-Free Protein Synthesis
作者:Isaac N. Arthur、James E. Hennessy、Dharshana Padmakshan、Dannon J. Stigers、Stéphanie Lesturgez、Samuel A. Fraser、Mantas Liutkus、Gottfried Otting、John G. Oakeshott、Christopher J. Easton
DOI:10.1002/chem.201203923
日期:2013.5.17
(DE3) used for cell‐free protein synthesis removes a wide range of α‐amino acid protecting groups by cleaving α‐carboxyl hydrazides; methyl, benzyl, tert‐butyl, and adamantyl esters; tert‐butyl and adamantyl carboxamides; α‐amino form‐, acet‐, trifluoroacet‐, and benzamides; and side‐chain hydrazides and esters. The free amino acids are produced and incorporated into a protein under standard conditions
大肠杆菌BL21 Star(DE3)的S30提取物可用于无细胞蛋白质合成,可通过裂解α-羧基酰肼去除大量的α-氨基酸保护基团。甲基,苄基,叔丁基和金刚烷基酯; 叔丁基和金刚烷酰胺; α-氨基甲酰胺,乙酰胺,三氟乙酰胺和苯甲酰胺; 以及侧链酰肼和酯。在标准条件下产生游离氨基酸并将其掺入蛋白质中。该方法允许在原位进行氨基酸的脱保护以避免单独的加工步骤。这种方法的优势通过有效结合化学难治性(S)-4-氟亮氨酸(S)-4,5-脱氢亮氨酸和(2 S,3 R)-4-氯缬氨酸通过直接使用它们各自的前体即(S)-4-氟亮氨酸酰肼(S)-4,5转变成蛋白质脱氢亮氨酸酰肼和(2 S,3 R)-4-氯缬氨酸甲酯。这些结果还表明,氟,脱氢亮氨酸和氯缬氨酸通过正常的生物合成机制被掺入蛋白质中,分别代替亮氨酸和异亮氨酸。