Method for improving protein synthesis efficiency in cells

Abstract

The invention provides a method for improving protein synthesis efficiency in cells. Specifically, the present invention provides an engineering bacterial strain for in-vitro cell-free protein synthesis, a first exogenous gene expression cassette of a first nucleic acid construct expressing a first fusion protein is integrated into a genome of the gene engineering bacterial strain, in the gene engineering bacterial strain, the expression or activity of a KlEXN53 gene (nuclease gene) is lowered. A cell extract (such as a yeast cell extract) derived from the engineering bacterial strain of the invention can significantly increase the stability of nucleic acid without the need for additional manual addition of T7 RNP, and the production efficiency of protein in an in-vitro protein synthesis system can be significantly increase.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method for improving protein synthesis efficiency in cells. Background technique [0002] Proteins are important molecules in cells and are involved in almost all functions of cells. Different sequences and structures of proteins determine their different functions. In cells, proteins can be used as enzymes to catalyze various biochemical reactions, and as signal molecules to coordinate various activities of organisms, to support biological forms, store energy, transport molecules, and make organisms move. In the field of biomedicine, protein antibodies, as targeted drugs, are an important means of treating diseases such as cancer [1-2]. [0003] In cells, protein production involves two parts: gene transcription and mRNA translation. [0004] Gene transcription refers to the process of using a strand of DNA as a template, under the catalysis of DNA-dependent RNA polymerase, and...

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Problems solved by technology

However, this system is not suitable for large-scale application due to adenosine triphosphate (ATP) imbalance [6]

Welch and Scopes solved the above problems through various explorations in 1985, and obtained high-yield ethanol, but this system also has two major defects: it needs to add high-cost enzymes and cannot tolerate temperature changes[7]

However, there are some inherent and difficult problems in this technology at present: such as reversibility, instability, leakage, inactivation, recycling of enzymes, lack of stable enzymes, enzyme complexes and cofactors, etc.

For example, only one sigma factor is present in cell-free extracts and transcriptional modularity remains poor

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