Yan Fang, Xi Chen, Zhen Sun, Xiaodan Yan, Lani Shi, Congcong Jin
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Discovery and investigation of the truncation of the (GGGGS)n linker and its effect on the productivity of bispecific antibodies expressed in mammalian cells.
Protein engineering is a powerful tool for designing or modifying therapeutic proteins for enhanced efficacy, increased safety, reduced immunogenicity, and improved delivery. Fusion proteins are an important group of therapeutic compounds that often require an ideal linker to combine diverse domains to fulfill the desired function. GGGGS [(G4S)n] linkers are commonly used during the engineering of proteins because of their flexibility and resistance to proteases. However, unexpected truncation was observed in the linker of a bispecific antibody, which presented challenges in terms of production and quality. In this work, a bispecific antibody containing 5*G4S was investigated, and the truncation position of the linkers was confirmed. Our investigation revealed that codon optimization, which can overcome the negative influence of a high repetition rate and high GC content in the (G4S)n linker, may reduce the truncation rate from 5-10% to 1-5%. Moreover, the probability of truncation when a shortened 3* or 4*G4S linker was used was much lower than that when a 5*G4S linker was used in mammalian cells. In the case of expressing a bispecific antibody, the bioactivity and purity of the product containing a shorter G4S linker were further investigated and are discussed.
期刊介绍:
Bioprocess and Biosystems Engineering provides an international peer-reviewed forum to facilitate the discussion between engineering and biological science to find efficient solutions in the development and improvement of bioprocesses. The aim of the journal is to focus more attention on the multidisciplinary approaches for integrative bioprocess design. Of special interest are the rational manipulation of biosystems through metabolic engineering techniques to provide new biocatalysts as well as the model based design of bioprocesses (up-stream processing, bioreactor operation and downstream processing) that will lead to new and sustainable production processes.
Contributions are targeted at new approaches for rational and evolutive design of cellular systems by taking into account the environment and constraints of technical production processes, integration of recombinant technology and process design, as well as new hybrid intersections such as bioinformatics and process systems engineering. Manuscripts concerning the design, simulation, experimental validation, control, and economic as well as ecological evaluation of novel processes using biosystems or parts thereof (e.g., enzymes, microorganisms, mammalian cells, plant cells, or tissue), their related products, or technical devices are also encouraged.
The Editors will consider papers for publication based on novelty, their impact on biotechnological production and their contribution to the advancement of bioprocess and biosystems engineering science. Submission of papers dealing with routine aspects of bioprocess engineering (e.g., routine application of established methodologies, and description of established equipment) are discouraged.