Affinity-Directed Site-Specific Protein Labeling and Its Application to Antibody-Drug Conjugates.

Chemically modified proteins have diverse applications; however, conventional chemo-selective methods often yield heterogeneously labeled products. To address this limitation, site-specific protein labeling holds significant potential, driving extensive research in this area. Nevertheless, site-specific modification of native proteins remains challenging owing to the complexity of their functional groups. Therefore, a method for site-selective labeling of intact proteins is aimed to design. In this study, a novel approach to traceless affinity-directed intact protein labeling is established, which leverages small binding proteins and genetic code expansion technology. By applying this method, a site-specific antibody labeling with a drug, which leads to the production of highly effective antibody-drug conjugates specifically targeting breast cancer cell lines is achieved. This approach enables traceless conjugation of intact target proteins, which is a critical advantage in pharmaceutical applications. Furthermore, small helical binding proteins can be easily engineered for various target proteins, thereby expanding their potential applications in diverse fields. This innovative approach represents a significant advancement in site-specific modification of native proteins, including antibodies. It also bears immense potential for facilitating the development of therapeutic agents for various diseases.

Engineering Translation Components for Genetic Code Expansion.

The expansion of the genetic code consisting of four bases and 20 amino acids into diverse building blocks has been an exciting topic in synthetic biology. Many biochemical components are involved in gene expression; therefore, adding a new component to the genetic code requires engineering many other components that interact with it. Genetic code expansion has advanced significantly for the last two decades with the engineering of several components involved in protein synthesis. These components include tRNA/aminoacyl-tRNA synthetase, new codons, ribosomes, and elongation factor Tu. In addition, biosynthesis and enhanced uptake of non-canonical amino acids have been attempted and have made meaningful progress. This review discusses the efforts to engineer these translation components, to improve the genetic code expansion technology.