Burkhart Brandon J, Kakkar Nidhi, Hudson Graham A, van der Donk Wilfred A, Mitchell Douglas A
Department of Chemistry, University of Illinois at Urbana-Champaign, 600 South Mathews Avenue, Urbana, Illinois 61801, United States.
Carl R. Woese Institute for Genomic Biology, University of Illinois at Urbana-Champaign, 1206 West Gregory Drive, Urbana, Illinois 61801, United States.
ACS Cent Sci. 2017 Jun 28;3(6):629-638. doi: 10.1021/acscentsci.7b00141. Epub 2017 Jun 6.
Combining biosynthetic enzymes from multiple pathways is an attractive approach for producing molecules with desired structural features; however, progress has been hampered by the incompatibility of enzymes from unrelated pathways and intolerance toward alternative substrates. Ribosomally synthesized and posttranslationally modified peptides (RiPPs) are a diverse natural product class that employs a biosynthetic logic that is highly amenable to engineering new compounds. RiPP biosynthetic proteins modify their substrates by binding to a motif typically located in the N-terminal leader region of the precursor peptide. Here, we exploit this feature by designing leader peptides that enable recognition and processing by multiple enzymes from unrelated RiPP pathways. Using this broadly applicable strategy, a thiazoline-forming cyclodehydratase was combined with enzymes from the sactipeptide and lanthipeptide families to create new-to-nature hybrid RiPPs. We also provide insight into design features that enable control over the hybrid biosynthesis to optimize enzyme compatibility and establish a general platform for engineering additional hybrid RiPPs.
组合来自多个途径的生物合成酶是生产具有所需结构特征分子的一种有吸引力的方法；然而，来自不相关途径的酶之间的不相容性以及对替代底物的不耐受性阻碍了这一进展。核糖体合成和翻译后修饰肽（RiPPs）是一类多样的天然产物，其采用的生物合成逻辑非常适合用于设计新化合物。RiPP生物合成蛋白通过与通常位于前体肽N端前导区的基序结合来修饰其底物。在这里，我们通过设计能被来自不相关RiPP途径的多种酶识别和加工的前导肽来利用这一特性。使用这种广泛适用的策略，将一种形成噻唑啉的环脱水酶与来自硫肽和羊毛硫肽家族的酶组合，以创造新型的天然杂交RiPPs。我们还深入了解了能够控制杂交生物合成以优化酶兼容性的设计特征，并建立了一个用于设计更多杂交RiPPs的通用平台。
