An in Vitro Reconstitution System Defines the Defective Step in the Biogenesis of Mutated β-Actin Proteins.

In vitro reconstitution of whole cellular events is one of the important goals in synthetic biology. Using a cell-free protein synthesis (CFPS) system reconstituted with human translation factors and chaperones, we reproduced the biogenesis of β-actin, synthesis, folding, and polymerization in a test tube. This system enabled us to define which step of the β-actin biogenesis was defective in genetic mutations related to diseases. Hence, the CFPS system reconstituted with human factors may be a useful tool for analyzing proteostasis in eukaryotes.