Protein production device and method

Abstract

The invention discloses a protein production device and method, the protein production device comprises a reaction chamber, a supplement chamber and a dialysis membrane, the reaction chamber is communicated with a supply device of a liquid-phase cell-free protein synthesis reaction system, and the supplement chamber is communicated with a supply device of a liquid-phase material supplement system; the dialysis membrane is used for separating the reaction chamber from the supplement chamber, and by-products which are generated in the liquid-phase cell-free protein synthesis reaction system and are used for inhibiting protein synthesis reaction and waste gas such as carbon dioxide can continuously penetrate through the dialysis membrane to be removed; in addition, a substrate required by the protein synthesis reaction in the liquid-phase material supplement system can continuously penetrate through the dialysis membrane to be supplemented into the reaction unit. According to the protein production device and method provided by the invention, a continuous fed-batch mode and a continuous exchange mode of cell-free protein production are combined, so that the overall size of a reaction container is effectively reduced, and the reaction is more flexible.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a protein production device and method. Background technique [0002] In recent years, cell-free protein synthesis (CFPS), as a new protein synthesis method, has shown great potential and application prospects. The main components of this protein synthesis system include crude extracts with basic transcription and translation functions, DNA templates, energy regeneration substances, cofactors and inorganic salts, etc. Different from protein synthesis by living cells in the natural state, the cell-free protein synthesis system is not restricted by the cell membrane, which not only allows the cell-free protein system to synthesize proteins that are harmful to cell activity, but also makes it easier for genes to be edited, and the protein synthesis process is better monitored. Greatly increases the flexibility of the protein synthesis process. Therefore, cell-free synthesi...

AI Technical Summary

Problems solved by technology

[0007] However, the existing cell-free expression system is difficult to achieve large-scale application of proteins in terms of cost and yield, and the existing problems are as follows:

[0008] (1) The batch reaction (batch) system is closed. During the system reaction, the required substances cannot be added. As the reaction proceeds, energy and substrates are gradually consumed, and metabolites and free phosphates accumulate. Inhibiting the reaction leads to a short lifespan of the system. The minimum protein expression system is mostly 50 μL and the reaction time is about 3 hours. The protein yield is low, usually only reaching μg mL-1

[0009] (2) Continuous-flow production mode In the reaction process, it is necessary to continuously add the substances and energy required for the reaction, and regularly remove the reaction by-products. The cumbersome operation of continuous-flow and the complexity of the device are serious. limits the usefulness of the model

[0011] The existing continuous exchange cell-free production methods such as figure 1 As shown, in this system, the reaction mixture in the dialysis tube cannot be mixed or stirred, the reaction rate is relatively low, there is a certain volume requirement, and the conditions of the reaction system are more stringent; Heat will be generated when stirred in a container, and the temperature of the overall reaction system is uncontrollable, which is not suitable for some temperature-sensitive synthetic reactions, and the dialysis tube cannot be used twice, and the price of the dialysis tube is relatively expensive, which increases the cost per experiment

[0012] In addition, the existing technology does not solve the aeration problem during the reaction process, and oxygen, as a key component in the regeneration system of adenosine triphosphate (ATP), has an important impact on the regeneration process of ATP

Insufficient oxygen supply reduces the rate of ATP regeneration and increases deoxyribonucleic acid (DNA) transcription-translation time, thereby reducing protein production and synthesis efficiency

At the same time, carbon dioxide produced during protein synthesis may also inhibit metabolic processes and affect protein expression

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