Galactosidase

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Galactosidase refers to a class of enzymes that hydrolyze substances containing galactosidic bonds, such as lactose (lactose is a disaccharide formed by dehydration and condensation of one molecule of glucose and one molecule of galactose). Mainly divided into α-galactosidase and β-galactosidase. α-Galactosidase catalyzes the hydrolysis of α-galactosidic bonds, which can transform and decompose the anti-nutritional factors α-galactosides in feed and soybean foods, and improve their nutritional content. In addition, the enzyme also has certain applications in pharmacy, thickener treatment and paper industry. β-Galactosidase not only has more and more uses in the food industry, but also plays an important role in biotechnology fields such as genetic engineering, enzyme engineering, protein engineering, etc., and has begun to be widely used in medicine and other fields.

Features

α-galactosidaseis an exoglycosidase that catalyzes the hydrolysis of α-galactosidic bonds. It is also called melibiose because it can decompose melibiose. α-galactosidaseis can catalyze the hydrolysis of α-galactosidic bonds. This feature makes it can be used to improve and eliminate anti-nutritional ingredients in feed and soy foods. In addition, it can achieve B→O blood type conversion in the medical field, prepare universal blood, and also play an important role in the enzyme replacement therapy of Fabry disease. α-Galactosidase can also act on complex polysaccharides, glycoproteins and glycosphingoses containing α-galactosidic bonds. Certain α-galactosidase enzymes also have the function of transgalactosylation when the substrate concentration is highly enriched. This feature can be used for the synthesis of oligosaccharides and the preparation of cyclodextrin derivatives. The development of neutrophilic or pH-stable α-galactosidase and the search for microorganisms or plants with high enzyme production have become research hotspots in recent years. Many heat-resistant alpha-galactosidase enzymes have gradually attracted wide interest from scientists due to their particularities. They expect to use their thermal stability to play a greater value in industry, as well as to show a wider range of technology.

β-galactosidase

β-galactosidase is a glycoside hydrolase, and there are many microbial sources. In addition to hydrolytic activity, some sources of β-galactosidase also have transglycosyl activity. Enzymes from different sources have different characteristics, such as the optimum pH, optimum temperature, and kinetic constant Km of the enzyme. In addition, the sources of enzymes are different, such as Kluyveromyces, Aspergillus, Bacillus, Streptococcus and Cryptococcus, and different reaction conditions, such as temperature and pH, the degree of polymerization, yield and formation of galactooligosaccharides synthesized by the enzyme reaction. The types of glycosidic bonds are also different.

Figure 1. Protein structure of β-galactosidase.

Mechanism

β-galactosidase can not only catalyze the hydrolysis of β-galactosidic bonds in β-galactosidic compounds, but also has the effect of transgalactosyl. Early studies have shown that the active site on β-galactosidase has two functional groups: Cys sulfhydryl and His imidazole, which play an important role in the hydrolysis of lactose by β-galactosidase. It is speculated that the sulfur group can be used as pantothenic acid to protonate the oxygen atom of galactosides, while the imidazole group can be used as a nucleophile to attack the nucleophilic center on the first carbon atom of the galactose molecule to form a carbon atom. After being cleaved off the imidazole group, the sulfhydryl anion extracts a proton from the water molecule to form -OH and attack C. Some studies believe that the catalytic mechanism of lactase is similar to that of lysozyme.

Application as a reporter gene

β-galactosidase gene is the most commonly used and most mature reporter gene in animal and microbial genetic engineering. A reporter gene is a gene that encodes a detectable protein or enzyme, and its phenotype should be easy to detect and distinguish from endogenous background proteins. Use its expression product to calibrate the expression regulation of the target gene. The application of β-galactosidase gene as a reporter gene mainly includes the following aspects: ① It is used to study the efficiency of promoters and the effect of mutations at different sites of the promoter on expression efficiency; ② It is used to study the enhancement sequence in expression systems, etc. The function of the regulatory sequence; ③ used to measure the expression characteristics of the vector and the influence of foreign substances on expression regulation; ④ used in the form of fusion genes to study the expression of foreign genes and their laws.