Since enzymes have the advantages of reaction specificity, high catalytic efficiency and mild reaction conditions, they have been increasingly used in industry, agriculture, medicine and environmental protection. But the overall has not yet reached the level of large-scale application, the main reason is that the enzyme itself has some deficiencies, such as instability, strict requirements for pH, and antigenicity. Therefore, people hope to use various methods to directionally modify enzyme molecules as needed, and even create new enzymes that have not yet been discovered in nature to meet the needs of all walks of life.
Method for transforming enzyme molecule
At present, there are two main methods for transforming enzyme molecules, namely chemical modification and biological
enzyme engineering.
Chemical modification uses chemical methods to transform enzyme molecules. The technique of chemical modification of enzyme molecules is relatively simple, but after modification of most enzymes, the chemical and biological properties will change. Therefore, the modification method should be selected according to the specific situation, and some protective measures should be taken to maintain the stability and yield of the enzyme as much as possible.
Biological enzyme engineering method: The chemical modification of enzymes is not the only way to transform enzymes. With the in-depth study of enzymes, the determination of the primary structure of amino acids, the application of gene recombination technology, etc., enzymes can be completely transformed, synthesized, and simulated. This is the main content of biological enzyme engineering. Biological enzyme engineering mainly includes genetic engineering technology to produce enzymes and protein engineering technology to transform enzymes.
Gene engineering technology to produce enzymes: Since the establishment of recombinant DNA technology in the 1970s, people have largely shed their dependence on natural enzymes. The development of genetic engineering makes it easier for people to obtain many kinds of natural enzyme genes by cloning, and make them efficiently expressed in microorganisms, and then carry out large-scale production through fermentation technology. Using genetic engineering methods to produce enzymes can greatly reduce the cost of enzyme products, and at the same time can make the production of rare enzymes easier.
At present, more than 100 enzyme genes have been successfully cloned, including urokinase gene and rennet gene. Among them, plasminogen activator and chymotrypsin are the most successful examples of using genetic engineering to obtain a large number of enzymes.
Human plasminogen activator is a class of serine
proteases. It can hydrolyze plasminogen to produce active plasmin and dissolve fibrin in the blood clot. Clinically used to treat thrombotic diseases and promote the dissolution of thrombus in the body. The plasminogen activator produced by the engineering strain has the same effect as the enzyme synthesized by the human body in curative effect, and has been used in clinical trials.
Chymotrypsin is an essential enzyme for cheese production, and its source is limited. Chymotrypsin extracted from microorganisms often causes cheese bitterness. Therefore, people cloned the calf rennet gene and expressed it in the yeast system, and the obtained rennet had completely the same properties as the natural enzyme extracted from the calf stomach.
Protein engineering technology to transform enzymes was developed on the basis of genetic engineering. The two have different contributions to enzyme engineering. Genetic engineering mainly solves the problem of mass production of enzymes, while protein engineering mainly modifies (or transforms) natural proteins to obtain enzymes with new functions or create completely new enzyme molecules.
Protein engineering enzymes change or remove certain amino acid residues in the primary structure of the enzyme protein, thereby changing the enzyme-related functions, so that the enzyme exhibits some new characteristics without affecting other functions.
At present, enzyme protein engineering is mainly used for the transformation of industrial enzymes. For example, Bacillus subtilis protease added to washing powder to enhance the detergency, but this enzyme is easy to lose activity under the action of bleach. Now the protein engineering technology is used to replace the amino acid in the enzyme protein molecule, which greatly improves the antioxidant capacity of the enzyme, so it can be used together with bleach.
Using protein engineering can also design new enzymes. From the current level of protein engineering development, it is still difficult to design completely new enzymes. However, with the in-depth study of protein chemistry, protein crystallography, enzymology and other related disciplines, the development of protein engineering will inevitably lead to a higher level.