Lysozyme is an enzyme present in both animal and human lacrimal gland secretions (or tears), gastric secretions, nasal mucus, and egg white. It is discovered in 1921 by Sir Alexander Fleming. The lysozymes catalyze the breakdown of certain carbohydrates that are found in the cell walls of certain bacteria (for example, cocci). As a result, in the case of lacrimal fluid, it protects the cornea of the eye from infection.
Lysozyme is most effective against the bacteria of Gram positive.
Structure of Lysozyme
A single peptide chain of around 129 amino acids makes up the condensed structure of lysozyme from hen egg white (hen egg white lysozyme or egg lysozyme). The residues of amino acid are numbered from the terminal α-group (N) to the terminal carboxyl-group (C). The circles show every fifth and every tenth residue is numbered. And, the broken lines indicate the four disulfide bridges. In the ranges of 25 to 35, 90 to 100, and 120 to 125, alpha-helices can be seen.
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Lysozyme crystals are stained with the methylene blue as shown below:
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T7 lysozyme or Bacteriophage and T4 are some of the examples of lysozyme.
The lysozyme enzyme breaks glycosidic bonds in peptidoglycans by hydrolyzing, attacking, and breaking them. Also, this particular lysozyme enzyme can break glycosidic bonds in chitin, although not as effective as true chitinases.
In the prominent cleft between its two domains, the Lysozyme's active site binds the peptidoglycan molecule. Between the N-acetylmuramic acid (NAM) and the N-acetylglucosamine (NAG) fourth carbon atom, it attacks the peptidoglycans (which are found in the cell walls of bacteria, especially Gram-positive bacteria).
Shorter saccharides such as tetrasaccharide have also been represented to be viable substrates but via an intermediate with a very long chain. Also, chitin has been determined to be one of the viable lysozyme substrates. And, artificial substrates have been developed and can be used in lysozyme.
The Phillips Mechanism has proposed that the catalytic power of the enzyme came from both steric strains on the electrostatic stabilization and bound substrate of an oxo-carbenium intermediate. From the data of X-ray crystallographic, Phillips has proposed the enzyme's active site, where a hexasaccharide binds. Also, lysozyme distorts the fourth-sugar (either in the D or -1 subsite) in the hexasaccharide into a conformation of half-chair.
In this particular stressed state, the glycosidic bond is broken very easily. An ionic intermediate having an oxo-carbenium can be created as a result of glycosidic bond breaking. As a result, the distortion lowers the reaction's energy barrier by forcing the substrate molecule to adopt a strained conformation close to that of the transition state.
Dahlquist suggested a covalent mechanism for lysozyme in 1969, based on the kinetic isotope effect, but the ionic mechanism was more widely accepted for a long time. In 2001, one revised mechanism was proposed by chemist Vocadlo via a covalent, but it is not an ionic intermediate. At the same time, the evidence from ESI-MS analysis has indicated a covalent intermediate. To lower the reaction and accumulate an intermediate for characterization, a 2-fluoro substituted substrate was used. This enzyme's activity is dependent on the amino acid side chains aspartate 52 (Asp52) and glutamic acid 35 (Glu35).
Imidazole derivatives may form the charge-transfer complex with a few residues (either in or outside the active center) to achieve a lysozyme's competitive inhibition. Lipopolysaccharide serves as a non-competitive inhibitor in Gram-negative bacteria by forming a strong bond with the lysozyme.
Even though the function of the muramidase lysozyme is thought to play a key role in its antibacterial properties, its non-enzymatic action has been identified as well. Blocking the catalytic activity of lysozyme by changing an essential amino acid in the active site (52-Asp -> 52-Ser), for example, does not render it antimicrobial. For tetrasaccharides similar to Klebsiella pneumonia lipopolysaccharide, lysozyme was found to have lectin-like ability to recognise the bacterial carbohydrate antigen without lytic operation. Also, the lysozyme interacts with T-cell receptors and antibodies.
Enzyme Conformation Changes
Lysozyme presents two conformations: an open, active state and a closed inactive state. The catalytic significance was investigated using single-walled carbon nanotubes (SWCN) field-effect transistors (FETs), which were attached to a single lysozyme. Electronically monitoring the lysozyme exhibited two conformations, where one is an open, active site and the other is a closed inactive site.
In the active state, lysozyme can be able to processively hydrolyze its substrate by breaking on the average 100 bonds at a rate of 15/sec. The active state requires two conformation phase changes to bind a new substrate and to transition from the closed inactive state to the open state, while the inactive state only requires one.
The other usable materials for biomedical and catalysis applications have been grown using lysozyme crystals. Commonly, the use of lysozyme can be implemented for lysing gram-positive bacteria. Lysozyme is widely used in the lab setting to release proteins from bacterium periplasm while the inner membrane remains sealed as vesicles known as spheroplast, due to its specific role of digesting the cell wall and causing an osmotic shock (burst the cell by changing the solute concentration unexpectedly around the cell and hence the osmotic pressure).