Host bacteria that have been genetically modified have a long history of producing certain proteins, as well as single enzymes for the alteration of chemicals made for industrial use by biological or chemical methods. These procedures have mostly been created through the process of discovery. The ability to design and assemble suitable enzymes into pathways that may or may not occur in nature to give a high-impact platform for the bio-manufacturing of chemicals, biofuels, and pharmaceuticals is a new development that is made possible by the synthetic biology method. Since whole cell synthesis is difficult or impossible to manage, a cell-free bio catalysis enables the manipulation of substrate ratios, the provision of regenerated cofactors, and the modification of high energy flux ratios. Here, we talk about the creation of cell-free bio catalytic pathways that may include additional free enzymes or multi-enzyme modules with heterologous catalysts. We review the state of commercializationrelated applications while highlighting the significance of economical cofactor regeneration. However, issues still exist, especially with regards to post-translational changes of proteins, such as glycosylation, phosphorylation, ubiquitination, acetylation, and proteolysis. Lists of the top value-added compounds from biomass, including glucaric acid, have been published by the National Renewable Energy Laboratory and Pacific North West Laboratory. There aren't many publications on how to create pathways for these useful intermediate chemicals using synthetic biology and cell-free protein synthesis. Despite at least one dedicated large-scale cell-free manufacturing of antibody conjugates, this observation remains. This review will outline one successful attempt at producing glucaric acid without the use of cells and assess advancements for additional important intermediate and platform chemicals.
