Translation Related Factors Improve the Productivity of a <i>Streptomyces</i>-Based Cell-Free Protein Synthesis System

Abstract

Cell-free protein synthesis (CFPS) systems are emerging as effective platforms for the production of recombinant proteins <i>in vitro</i>. To enable the expression of various proteins, different CFPS systems have been developed to better mimic the cellular environment of native hosts. In this context, a <i>Streptomyces</i>-based CFPS system was recently developed to express high GC-content genes. Unfortunately, protein yields from the initial system were relatively low (∼50 μg/mL). Here, we sought to address this limitation and enhance the productivity of the <i>Streptomyces</i>-based CFPS system. By adding protein translation related factors to CFPS reactions, we were able to achieve protein yields of approximately 400 μg/mL, which is the highest yield reported to date. We expect our enhanced <i>Streptomyces</i> CFPS system will set the stage for novel applications in metabolic engineering and synthetic biology such as <i>in vitro</i> discovery and synthesis of natural products, which are produced by <i>Streptomyces</i> species with high GC-content (>70%) genomes.