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Lysozyme (, muramidase, N-acetylmuramide glycanhydrolase; systematic name peptidoglycan N-acetylmuramoylhydrolase) is an antimicrobial enzyme produced by animals that forms part of the innate immune system. It is a glycoside hydrolase that catalyzes the following process:
Peptidoglycan is the major component of gram-positive bacterial cell wall. This hydrolysis in turn compromises the integrity of bacterial cell walls causing lysis of the bacteria.
Lysozyme is abundant in including tears, saliva, human milk, and mucus. It is also present in cytoplasmic granules of the macrophages and the polymorphonuclear neutrophils (PMNs). Large amounts of lysozyme can be found in egg white. C-type lysozymes are closely related to α-lactalbumin in sequence and structure, making them part of the same glycoside hydrolase family 22. In humans, the C-type lysozyme enzyme is encoded by the LYZ gene.
Hen egg white lysozyme is thermally stable, with a melting point reaching up to 72 °C at pH 5.0. However, lysozyme in human milk loses activity very quickly at that temperature. Hen egg white lysozyme maintains its activity in a large range of pH (6–9). Its isoelectric point is 11.35. The isoelectric point of human milk lysozyme is 10.5–11.
's active site binds the peptidoglycan molecule in the prominent cleft between its two domains. It attacks peptidoglycans (found in the cell walls of bacteria, especially Gram-positive bacteria), its natural substrate, between N-acetylmuramic acid (NAM) and the fourth carbon atom of N-acetylglucosamine (NAG).
Shorter Carbohydrate like tetrasaccharide have also shown to be viable substrates but via an intermediate with a longer chain. Chitin has also been shown to be a viable lysozyme substrate. Artificial substrates have also been developed and used in lysozyme. (1996). 9783034899529 ISBN 9783034899529
The proposed oxo-carbonium intermediate was speculated to be electrostatically stabilized by aspartate and glutamate residues in the active site by Arieh Warshel in 1978. The electrostatic stabilization argument was based on comparison to bulk water, the reorientation of water dipoles can cancel out the stabilizing energy of charge interaction. In Warshel's model, the enzyme acts as a super-solvent, which fixes the orientation of ion pairs and provides super-solvation (very good stabilization of ion pairs), and especially lower the energy when two ions are close to each other.
The rate-determining step (RDS) in this mechanism is related to formation of the Oxocarbenium intermediate. There were some contradictory results to indicate the exact RDS. By tracing the formation of product (p-nitrophenol), it was discovered that the RDS can change over different temperatures, which was a reason for those contradictory results. At a higher temperature the RDS is formation of glycosyl enzyme intermediate and at a lower temperature the breakdown of that intermediate.
More recently, quantum mechanics/ molecular mechanics (QM/MM) molecular dynamics simulations have been using the crystal of HEWL and predict the existence of a covalent intermediate. Evidence for the ESI-MS and X-ray structures indicate the existence of covalent intermediate, but primarily rely on using a less active mutant or non-native substrate. Thus, QM/MM molecular dynamics provides the unique ability to directly investigate the mechanism of wild-type HEWL and native substrate. The calculations revealed that the covalent intermediate from the covalent mechanism is ~30 kcal/mol more stable than the ionic intermediate from the Phillips mechanism. These calculations demonstrate that the ionic intermediate is extremely energetically unfavorable and the covalent intermediates observed from experiments using less active mutant or non-native substrates provide useful insight into the mechanism of wild-type HEWL.
Since lysozyme is a natural form of protection from Gram-positive pathogens like Bacillus and Streptococcus, (2025). 9780071107068, McGraw-Hill Higher Education. ISBN 9780071107068 it plays an important role in immunology of infants in human milk feeding. Whereas the skin is a protective barrier due to its dryness and acidity, the conjunctiva (membrane covering the eye) is, instead, protected by secreted enzymes, mainly lysozyme and defensin. However, when these protective barriers fail, conjunctivitis results.
In certain cancers (especially myelomonocytic leukemia) excessive production of lysozyme by cancer cells can lead to toxic levels of lysozyme in the blood. High lysozyme blood levels can lead to kidney failure and low blood potassium, conditions that may improve or resolve with treatment of the primary malignancy.
Serum lysozyme is much less specific for diagnosis of sarcoidosis than serum angiotensin converting enzyme; however, since it is more sensitive, it is used as a marker of sarcoidosis disease activity and is suitable for disease monitoring in proven cases.
For example, E. coli can be lysed using lysozyme to free the contents of the space. It is especially useful in lab setting for trying to collect the contents of the periplasm. Lysozyme treatment is optimal at particular temperatures, pH ranges, and salt concentrations. Lysozyme activity increases with increasing temperatures, up to 60 degrees Celsius, with a pH range of 6.0-7.0. The salts present also affect lysozyme treatment, where some assert inhibitory effects, and others promote lysis via lysozyme treatment. Sodium chloride induces lysis, but at high concentrations, it is an active inhibitor of lysis. Similar observations have been seen with the use of potassium salts. Slight variations are present due to differences in bacterial strains. A consequence of the use of lysozyme in extracting recombinant proteins for protein crystallization is that the crystal may be contaminated with units of lysozyme, producing a physiologically irrelevant combination. In fact, some proteins simply cannot crystalize without such contamination.
Furthermore, lysozyme can serve as a tool in the expression of toxic recombinant proteins. Expressing recombinant proteins in BL21(DE3) strains is typically accomplished by the T7-RNA-polymerase. Via IPTG induction, the UV-5 repressor is inhibited, leading to the transcription of the T7-RNA-polymerase and thereby of the protein of interest. Nonetheless, a basal level of the T7-RNA-polymerase is observable even without induction. T7 lysozyme acts as an inhibitor of the T7-RNA-polymerase. Newly invented strains, containing a helper plasmid (pLysS), constitutively co-express low levels of T7 lysozyme, providing high stringency and consistent expression of the toxic recombinant protein.
Lysozyme was first crystallised by Edward Abraham in 1937, enabling the three-dimensional structure of hen egg white lysozyme to be described by David Chilton Phillips in 1965, when he obtained the first 2-ångström (200 picometer) resolution model via X-ray crystallography. The structure was publicly presented at a Royal Institution lecture in 1965. Lysozyme was the second protein structure and the first enzyme structure to be solved via X-ray diffraction methods, and the first enzyme to be fully sequenced that contains all twenty common amino acids. As a result of Phillips' elucidation of the structure of lysozyme, it was also the first enzyme to have a detailed, specific mechanism suggested for its method of catalytic action. This work led Phillips to provide an explanation for how enzymes speed up a chemical reaction in terms of its physical structures. The original mechanism proposed by Phillips was more recently revised.
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