Computational design and selections for an engineered, thermostable terpene synthase
Computational design and selections for an engineered, thermostable terpene synthase
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DOI：
10.1002/pro.691
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发表时间：
2011-09-01
期刊：
影响因子：
8
通讯作者：
Weiss, Gregory A.
中科院分区：
文献类型：
--
作者：
Diaz, Juan E.;Lin, Chun-Shi;Weiss, Gregory A.
萜类化合物包括结构多样的抗生素、调味剂和香料。工程萜类合成酶控制这些化合物的合成是一个长期追求的目标。我们报道了在高温下催化碳正离子环化反应的烟草5-马兜铃烯合成酶（TEAS）耐热突变体的计算设计、选择和测定。热稳定性的选择包括蛋白质水解消化，然后捕获完整的蛋白质。与野生型酶不同，突变型TEAS在65℃下保持酶活性。耐热萜烯合成酶变体在80℃以上变性，大约是野生型酶温度的两倍。
Terpenoids include structurally diverse antibiotics, flavorings, and fragrances. Engineering terpene synthases for control over the synthesis of such compounds represents a long sought goal. We report computational design, selections, and assays of a thermostable mutant of tobacco 5-epi-aristolochene synthase (TEAS) for the catalysis of carbocation cyclization reactions at elevated temperatures. Selection for thermostability included proteolytic digestion followed by capture of intact proteins. Unlike the wild-type enzyme, the mutant TEAS retains enzymatic activity at 65 degrees C. The thermostable terpene synthase variant denatures above 80 degrees C, approximately twice the temperature of the wild-type enzyme.