Smith Mason R, Gao Hui, Prabhu Ponnandy, Bugada Luke F, Roth Cori, Mutukuri Deepika, Yee Christine M, Lee Lester, Ziff Robert M, Lee Jung-Kul, Wen Fei
Department of Chemical Engineering, University of Michigan, Ann Arbor, Michigan 48109, United States.
Department of Chemical Engineering, Konkuk University, Seoul 05029, Republic of Korea.
Nat Catal. 2019;2(9):809-819. doi: 10.1038/s41929-019-0321-8. Epub 2019 Jul 22.
Cooperative enzyme catalysis in nature has long inspired the application of engineered multi-enzyme assemblies for industrial biocatalysis. Despite considerable interest, efforts to harness the activity of cell-surface displayed multi-enzyme assemblies have been based on trial and error rather than rational design due to a lack of quantitative tools. In this study, we developed a quantitative approach to whole-cell biocatalyst characterization enabling a comprehensive study of how yeast-surface displayed multi-enzyme assemblies form. Here we show that the multi-enzyme assembly efficiency is limited by molecular crowding on the yeast cell surface, and that maximizing enzyme density is the most important parameter for enhancing cellulose hydrolytic performance. Interestingly, we also observed that proximity effects are only synergistic when the average inter-enzyme distance is > ~130 nm. The findings and the quantitative approach developed in this work should help to advance the field of biocatalyst engineering from trial and error to rational design.
自然界中的协同酶催化长期以来一直启发着人们将工程化多酶组装体应用于工业生物催化。尽管备受关注，但由于缺乏定量工具，利用细胞表面展示的多酶组装体活性的努力一直基于试错法而非理性设计。在本研究中，我们开发了一种全细胞生物催化剂表征的定量方法，能够全面研究酵母表面展示的多酶组装体的形成方式。在此我们表明，多酶组装效率受到酵母细胞表面分子拥挤的限制，并且最大化酶密度是提高纤维素水解性能的最重要参数。有趣的是，我们还观察到，只有当平均酶间距离大于约130 nm时，邻近效应才具有协同作用。这项工作中得出的研究结果和开发的定量方法应有助于推动生物催化剂工程领域从试错法走向理性设计。
