Cell-Free Mixing of Escherichia coli Crude Extracts to Prototype and Rationally Engineer High-Titer Mevalonate Synthesis.

ABSTRACT: Cell-free metabolic engineering (CFME) is advancing a powerful paradigm for accelerating the design and synthesis of biosynthetic pathways. However, as most cell-free biomolecule synthesis systems to date use purified enzymes, energy and cofactor balance can be limiting. To address this challenge, we report a new CFME framework for building biosynthetic pathways by mixing multiple crude lysates, or extracts. In our modular approach, cell-free lysates, each selectively enriched with an overexpressed enzyme, are generated in parallel and then combinatorically mixed to construct a full biosynthetic pathway. Endogenous enzymes in the cell-free extract fuel high-level energy and cofactor regeneration. As a model, we apply our framework to synthesize mevalonate, an intermediate in isoprenoid syn

SUBMITTER: Dudley QM 

PROVIDER: S-EPMC6728267 | biostudies-literature | 2016 Dec

REPOSITORIES: biostudies-literature