Fittolani Giulio, Kutateladze Dennis A, Loas Andrei, Buchwald Stephen L, Pentelute Bradley L
Department of Chemistry, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, Massachusetts 02139, United States.
The Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, 500 Main Street, Cambridge, Massachusetts 02142, United States.
J Am Chem Soc. 2025 Feb 5;147(5):4188-4197. doi: 10.1021/jacs.4c13832. Epub 2025 Jan 22.
The remarkable efficiency with which enzymes catalyze small-molecule reactions has driven their widespread application in organic chemistry. Here, we employ automated fast-flow solid-phase synthesis to access catalytically active full-length enzymes without restrictions on the number and structure of noncanonical amino acids incorporated. We demonstrate the total syntheses of iron-dependent myoglobin (BsMb) and sperm whale myoglobin (SwMb). The synthetic enzymes displayed excellent enantioselectivity and yield in carbene transfer reactions. Absolute control over enantioselectivity in styrene cyclopropanation was achieved using synthetic L- and D-BsMb mutants, which delivered each enantiomer of cyclopropane product in identical and opposite enantiomeric enrichment. BsMb mutants outfitted with noncanonical amino acids were used to facilitate detailed structure-activity relationship studies, revealing a previously unrecognized hydrogen-bonding interaction as the primary driver of enantioselectivity in styrene cyclopropanation. We anticipate that our approach will advance biocatalysis by providing reliable and rapid access to fully synthetic enzymes possessing noncanonical amino acids.
酶催化小分子反应的卓越效率推动了它们在有机化学中的广泛应用。在此，我们采用自动化快速流动固相合成法来获取具有催化活性的全长酶，且对所掺入的非天然氨基酸的数量和结构没有限制。我们展示了铁依赖性肌红蛋白（BsMb）和抹香鲸肌红蛋白（SwMb）的全合成。合成酶在卡宾转移反应中表现出优异的对映选择性和产率。使用合成的L-和D-BsMb突变体实现了对苯乙烯环丙烷化中对映选择性的绝对控制，它们以相同但相反的对映体富集度提供环丙烷产物的每种对映体。配备非天然氨基酸的BsMb突变体用于促进详细的构效关系研究，揭示了一种先前未被认识到的氢键相互作用是苯乙烯环丙烷化中对映选择性的主要驱动力。我们预计，我们的方法将通过可靠且快速地获取具有非天然氨基酸的全合成酶来推动生物催化的发展。
