Lysozyme dimer

Lysozyme dimer The lysozyme dimer is a product of the process of polymerization of single molecules of natural Hen Egg-White lysozyme.


Lysozyme (muramidase) belongs to the class of enzymes that hydrolyze chemical bonds, known according to current systematic terminology as N-acetylmuramide glycan hydrolase. It was discovered in 1922 by Scottish physician and microbiologist Alexander Fleming, later winner of the Nobel Prize (1945). The enzyme is commonly found in nature. It can be isolated from the cells and body fluids of all animal species. That is why Fleming thought he had discovered a natural, filo genetically early stage, and primeval immune mechanism characteristic of all living organisms. In animals it occurs in several different forms, different for each taxonomic subtype: type c and type g in vertebrates, and type i (sometimes type c) in invertebrates. In the latter, for example in bees, it is one of the most important factors in natural immunity. As yet another example, the termite egg recognition pheromone, characteristic and relevant for advanced termite communities, is identical to lysozyme in its chemical composition and structure.



The spatial structure of a molecule of monomeric lysozyme ( X-ray structure pdb; Yikrazuul).



Along with research on lysozyme, Fleming intensively worked on his other groundbreaking discovery, penicillin. When he completed the first stage of research on penicillin and offered it for clinical use, he drove medicine into a completely different epoch, the era of antibiotics, with all their pros and cons.

At the time, the world stood on the verge of World War II and the new medication was soon to save thousands of human lives. The huge success of penicillin however, overshadowed Fleming’s first great achievement, the discovery of lysozyme. Lysozyme became known only in the second half of the XXth century. It was then that the enzyme became the object of intensive biochemical, physiological and clinical studies. Even though not all its properties have been identified, it is clear that lysozyme shows antibacterial, antiviral and anti-inflammatory activity.

The antibacterial activity is based on the catalyzing hydrolysis of 1,4-beta-linkages between N-acetylmuramic acid and N-acetyl-D-glucosamine, which occurs in the bacterial cell walls and may require the presence of or cooperation with other enzymes or antibodies. This explains its activity only in living organisms (in vivo).

Moreover, the lysozyme included in lysosomes is responsible for intracellular digestion in phagocytes. Many authors have confirmed the presence of lysozyme in phagocytes and its influence on phagocytosis. There are, however, relevant differences among species. For example, human lysozyme stimulates phagocytosis in physiological concentrations of 10-400 µg/ml, while the same enzyme isolated from the hen egg-white shows lower stimulation, which can be registered only at a concentration of 1000 µg/ml or higher.

The antiviral properties of the lysozyme are probably due to the increase of natural immunity and resemble the activity of interferon (INF). This activity is indirect and it stimulates a cell to produce INF type I (especially INFα) and type II (INFγ). Direct activity occurs through bonding of positively charged dimer molecules with negatively polarized nucleic acid chains, which form inactive units (compounds?) connected by electrostatic bonds. The lysozyme forms similar compounds with nucleoproteins and lipoproteins, which are essential to the development of viruses within cells.

Lysozyme also has anti-inflammatory activity. Due to its very alkaline properties, it neutralizes acid mediators of the inflammatory process and lowers body temperature when the process is caused by endogenic pyrogens.

Unfortunately, the application of preparations containing natural monomeric lysozyme did not bring the expected therapeutic results. These expectations were met only by the lysozyme dimer, which is a polypeptide with a molecular mass of about 27000 Daltons.

In natural conditions the lysozyme occurs probably exclusively as a monomer, but only at a certain pH, concentration and temperature. Depending on the changes of these parameters, single molecules of the enzyme bond into dimers and in more complex polymers. What changes then is not only the summary molecular mass or the volume of the new molecule, but also its biochemical properties (more»). From the medical point of view this is beneficial because the polymeric derivative retains the clinical activity of a monomer, but practically eliminates its negative properties, such as bio-toxicity. The molecule of the lysozyme dimer becomes spherical. This shape is a constant feature dependant on ionic strength and on pH and is therefore easy to describe by a simple physical formula. Dimerization is considered to be the first step in a reversible process of lysozyme crystallization, which can be carried out within laboratory conditions.

The process of bonding of monomeric molecules in nature is quite common and many natural substances become active only as polymers. Examples may be immunoglobulin A and kachectin, tumor necrosis factor (TNF), both of which occur in the dimer or trimer form. Immunoglobulin M occurs as a pentamer. Ribonuclease of bull semen and human interleukin IL-5 are both active dimers.

The NIKA HP Company has done extensive research in human and veterinary medicine on the application of the lysozyme dimer. This dimer was derived from the polymerization of the enzyme obtained from hen egg white. Our research was considered pioneering on a global scale (more»). These studies (in vivo on hens’ embryos) confirmed that the lysozyme dimer completely inhibits the proliferation of the Sendai virus at the relatively low dose of 0.01 mg/ml. It also reveals a strong inhibition towards the flu virus (virus A/Sichuan no 43/17), cowpox and measles. Therefore, it can be assumed to be effective in the fight against other viruses which have single stranded, negatively polarized, segmented RNA, as well as the recently described Schmallenberg virus and other orthobunyaviruses. It should again be emphasized here that the inhibiting effect of the lysozyme dimer on pathogens can be observed only in living organisms.

Comparative studies concerning the cytotoxicity of lysozyme on hen fibroblasts have proved the dimer to be considerably less toxic than its monomer. A dimer concentration as high as 1.0 mg/ml did not show any cytotoxicity up to the seventh day of cell culture. The dimer’s low toxicity was later confirmed on animals. The first symptoms of intolerance occurred only when a parenteral dose, higher than 200mg/ml body mass in mice (NMRI strain) and higher than 500-mg/kg body mass in rats, was administered. In dogs, intolerance was noticed only when a dose of 80 mg/kg body mass was used, i.e. 4000 times greater than the therapeutic dose.

One of the most interesting characteristics of the dimer is its ability to modulate the synthesis of kachectin (TNF), a polypeptide, which belongs to a group of cytokines and is produced by stimulated macrophages. It does not quite stunt the dimer’s synthesis, but only prevents its excessive production. Clinically this is beneficial against many diseases, e.g. acute diarrhea or cancer.

The application of the lysozyme dimer in veterinary practice has created many new possibilities in the prevention and treatment of various diseases, especially when used together with other medications, for example, antibiotics. The lysozyme dimer considerably decreases the negative immunosuppressive effect of antibiotics.

More on the lysozyme dimer in Publications.