Lysozyme

Lysozyme (EC 3.2.1.17) belongs to the class of glycoside hydrolases, is an alkaline enzyme that hydrolyzes mucopolysaccharides in pathogenic bacteria. The systematic name of this enzyme class is peptidoglycan N-acetylmuramoylhydrolase. Other names in common use include muramidase, globulin G, N-acetylmuramide glycanhydrolase and so on. Lysozyme was firstly identified in 1922, Alexander Fleming found an enzyme in human tears and saliva that can dissolve the cell wall of bacteria. Because of its lysozyme effect, he named it lysozyme. Since then, people have found lysozyme in various tissues and secretions of humans and animals, as well as certain plants and microorganisms. It is also possible to isolate lysozyme from plants such as fig, barley, and cabbage.

Classification

Lysozyme is divided into two major categories according to the microorganisms it acts on, including bacterial cell wall lysozyme and fungal cell wall lysozyme. Fungal cell wall lysozymes include yeast cell wall lysozyme and mold cell wall lysozyme. According to their different sources, they can be divided into animal source lysozyme, plant source lysozyme and microbial source lysozyme.

Structure

Lysozyme is the most well studied enzyme. The first to resolve the 3D structure is the structure of hen egg white lysozyme reported in 1965. It is a single peptide chain protein consisting of 129 amino acid residues, and 4 pairs of cysteines in the molecule make up 4 S-S bonds. The lysozyme molecule is elliptical and has a size of 4.5 nm×3.0 nm×3.0 nm. The molecule contains 25% of α-helix and some β-sheet structure. Studies have shown that the surface of the lysozyme molecule has a fissure that accommodates the polysaccharide substrate and is the active site of lysozyme. The interior of lysozyme is almost non-polar, and hydrophobic interactions play an important role in the folded conformation of lysozyme.

Figure 1. Structure of hen egg white lysozyme. (Held J. 2014)

Physicochemical properties

Lysozyme is a basic protein with an isoelectric point of 10.7~11.0. Its chemical properties are stable and its thermal stability is high. The optimum temperature for its activity is 45~50 °C. When the pH is between 1.2 and 11.3, the structure is almost unchanged. In an acidic environment, lysozyme has a high thermal stability. At pH 4~7, the original enzyme activity can be maintained at 100 °C for 1 min; at pH 3, it can be treated at 100 °C for 45 min. The egg white lysozyme has a molecular weight of 14000. The isoelectric point is 11.1, and the optimum bacteriostatic temperature is 50 °C. Optimum pH of this enzyme is 7. The original enzyme activity is maintained, but the enzyme is less thermally stable in an alkaline environment. Under dry conditions, lysozyme can be stored at room temperature for a long time. Its pure product is white or yellowish or yellow crystal or amorphous powder, odorless, sweet, soluble in water, easily destroyed by alkali, insoluble in acetone and ether, etc.

Catalytic Mechanism

The substrate of lysozyme is the peptidoglycan cell wall of gram-positive bacteria, which destroys the cell wall by cut the β(1→4) glycosidic bond between alternating units of N-acetylmuramic acid and N-acetylglucosamine. The active sites of the hen egg white lysozyme are Asp52 and Glu35. The terminal proton of Glu35 is transferred to the oxygen atom of the glycosidic bond between two adjacent sugar residues, thereby cutting of the glycosidic bond and producing of a carbenium ion. The positive charge of the carbenium ion is stabilized by the negative charge of Asp52 until a hydroxyl ion is bonded to the carbon positive atom, and then Glu35 is reprotonated.

Application

Lysozyme is now widely used in food and medicine. As a non-toxic, non-side-effect protein, lysozyme can be used as a natural food preservative. It has been widely used in the preservation of aquatic products, meat products, cakes, sake, cooking wine and beverages. It can also be added to milk powder to emulsify cow's milk to inhibit the survival of spoilage microorganisms in the intestine. As a non-specific immune factor existing in normal human body fluids and tissues, it has antibacterial, antiviral and antitumor effects that can be used as a natural anti-infective material. Besides, lysozyme has the function of destroying the structure of bacterial cell wall, and are used to deal with the G+ bacteria and obtain protoplasts. Therefore, lysozyme is an essential enzyme for cell fusion in genetic engineering and cell engineering.

References

1. Held J., Smaalen S.V. The active site of hen egg-white lysozyme: flexibility and chemical bonding. Acta Crystallographica, 2014, D70: 1136-1146.
2. Lin C.H., Xiao S.R, Meng Q.G. Structural characteristics of lysozyme and its application. Journal of Weifang University, 2005, 5(2).