为选择性四次碳键形成量身定制基合成酶TrpB
Markus Dick1, Nicholas S Sarai1, Michael W Martynowycz2
1Division of Chemistry and Chemical Engineering 210-41 , California Institute of Technology , 1200 East California Boulevard , Pasadena , California 91125 , United States.
Journal of the American Chemical Society
|November 21, 2019
概括
研究人员设计了一种酸合成酶变体,TrpB Pf_quat,以创建新的非正规氨基酸. 这种酶选择性地形成全碳四元立体中心,扩大合成化学能力.
科学领域:
- 生物催化
- 酶工程
- 有机合成
背景情况:
- 酸合成酶 (TrpB) 在自然情况下催化L-和凝到L-酸.
- 之前的工程工作使TrpB能够合成非正规的氨基酸.
- 在合成化学中创建四元立体中心仍然是一个挑战.
研究的目的:
- 设计TrpB以形成C-C键与3替代的氧化物.
- 开发一种能够产生新的四元立体中心的酶.
- 扩大工程TrpB用于合成各种非正规氨基酸的实用性.
主要方法:
- 在六代的时间里指导了TrpB酶的演变.
- 使用3替代氧作为基板,有或没有N1保护.
- 使用微晶电子衍射 (MicroED) 来确定立体中心配置.
主要成果:
- 改造的TrpB变体,TrpB Pf_quat,在3替代氧醇化中表现出400倍的活性增加.
- 该酶在氨基酸产品的玛位置选择性地形成全碳四元立体中心.
- TrpB Pf_quat显示了乳和中三级碳的区域选择性.
- 高酶热稳定性和表达水平 (大肠杆菌培养中的500mg/L).
结论:
- TrpB Pf_quat提供了一个高效和可持续的生物催化平台,用于构建非正规氨基酸与全碳四元立体中心.
- 这种工程酶扩大了生物催化剂中C-C键形成的范围.
- 微ED分析证实了新形成的立体中心的立体化学.
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