Nussbaumer Martin G, Nguyen Peter Q, Tay Pei K R, Naydich Alexander, Hysi Erisa, Botyanszki Zsofia, Joshi Neel S
Wyss Institute for Biologically Inspired Engineering, Harvard University, Boston, MA 02115 (USA).
Joshi School of Engineering and Applied Sciences Harvard University Cambridge, MA 02138 (USA).
ChemCatChem. 2017 Dec 8;9(23):4328-4333. doi: 10.1002/cctc.201701221. Epub 2017 Aug 2.
Cell-free biocatalysis systems offer many benefits for chemical manufacturing, but their widespread applicability is hindered by high costs associated with enzyme purification, modification, and immobilization on solid substrates, in addition to the cost of the material substrates themselves. Herein, we report a "bootstrapped" biocatalysis substrate material that is produced directly in bacterial culture and is derived from biofilm matrix proteins, which self-assemble into a nanofibrous mesh. We demonstrate that this material can simultaneously purify and immobilize multiple enzymes site specifically and directly from crude cell lysates by using a panel of genetically programmed, mutually orthogonal conjugation domains. We further demonstrate the utility of the technique in a bienzymatic stereoselective reduction coupled with a cofactor recycling scheme. The domains allow for several cycles of selective removal and replacement of enzymes under mild conditions to regenerate the catalyst system.
无细胞生物催化系统为化学制造带来诸多益处，但其广泛应用受到多种因素的阻碍，除了材料底物本身的成本外，还包括与酶纯化、修饰以及固定在固体底物上相关的高昂成本。在此，我们报告一种“自组装”的生物催化底物材料，它直接在细菌培养物中产生，源自生物膜基质蛋白，能自组装成纳米纤维网。我们证明，通过一组基因编程的、相互正交的共轭结构域，这种材料可以从粗细胞裂解物中位点特异性地直接同时纯化和固定多种酶。我们进一步展示了该技术在双酶立体选择性还原与辅因子循环方案相结合中的实用性。这些结构域允许在温和条件下进行几个循环的酶选择性去除和替换，以再生催化剂系统。
