Cell-Free PURE System:Evolution and Achievements

The cell-free protein synthesis (CFPS) system, as a technical core of synthetic biology, can simulate the transcription andtranslation process in an in vitro open environment without a complete living cell. It has been widely used in basic and appliedresearch fields because of its advanced engineering features in flexibility and controllability. Compared to a typical crudeextract-based CFPS system, due to defined and customizable components and lacking protein-degrading enzymes, the proteinsynthesis using recombinant elements (PURE) system draws great attention. This review first discusses the elementalcomposition of the PURE system. Then, the design and preparation of functional proteins for the PURE system, especially thecritical ribosome, were examined. Furthermore, we trace the evolving development of the PURE system in versatile areas,including prototyping, synthesis of unnatural proteins, peptides and complex proteins, and biosensors. Finally, as a state-ofthe-art engineering strategy, this review analyzes the opportunities and challenges faced by the PURE system in futurescientific research and diverse applications.