Attack the bacteria
Lysozyme protects us from the continuous danger of bacterial infections. It is a small enzyme that attacks the cell wall of bacteria. The bacteria have a robust coating of carbohydrate chains, transversely linked by small peptide chains, which wraps their delicate membrane to defend it from the strong internal osmotic pressure of the cell. Lysozyme breaks these carbohydrate chains, destroying the structural integrity of the cell wall and therefore the bacteria explode by their own internal pressure.
The first antibiotic
Alexander Fleming suppressed lysozyme while he was conducting targeted research to find antibiotic drugs. He continued for years to add everything that came to his mind to bacterial cultures, looking for something that would slow down their growth. Fleming discovered lysozyme by accident one day while he was having a severe cold, added a drop of mucus to a crop and to his surprise the bacteria died. He discovered one of our natural defenses against infections. Unfortunately, lysozyme is too large a molecule and cannot be used as a medicine. It can be applied topically, but it cannot rid the whole body of infections because it is too large to move between cells. Fortunately, Fleming continued his research until, five years later, he discovered a real antibiotic drug: penicillin (mdm 5/2002).
Lysozyme protects many places rich in potential food for the bacteria with which they could grow. The lysozyme shown above (PDB file 6jxp) was extracted from chicken egg white, where it protects the proteins and fats that feed the chick that is being formed. There are 4 disulfide bridges (with yellow sulfur atoms) which make the 3D structure of the protein more stable and, in the deep pocket of the active site where the carbohydrates that need to be cut bind, there are two amino acids of aspartic acid (red oxygen) .
Tears and mucus also contain lysozyme to fight infections on the most exposed surfaces. Blood is the most dangerous place where bacteria can grow, because they would be transported to all areas of the body. In the blood, lysozyme provides some protection, along with even more powerful weapons than the immune system.
Lysozyme is a small and stable enzyme that constitutes an ideal model for research on the structure and function of proteins.
Brian Matthews, at the University of Oregon, has performed a remarkable series of experiments using lysozyme as a molecular laboratory. He carried out hundreds of mutations on lysozyme using the bacteriophage technique, replacing one or more amino acids in the protein chain with other different ones. He studied the effects caused by the removal of bulky amino acids or the introduction of a large amino acid where normally there would not have been. He tried to create new active sites by creating new pockets elsewhere on the molecule.
The structures of hundreds of these mutant lysozymes are available in the PDB archives, in reality these structures are so numerous that lysozyme represents the most common protein in the PDB archives. The figure opposite on the right (file PDB 1l35) shows a mutant in which two amino acids (shown in green) have been replaced with cysteine, forming a new disulfide bridge (two yellow atoms). On the left, for comparison, the native enzyme is shown (PDB 1lyd file).
Exploring the structure
Lysozyme has a long-slit-shaped active site that binds to the carbohydrate chains of the bacteria cell wall. The structure shown here (file PDB 148l) contains a fragment of a bacterial cell wall, with two sugars with a cyclic structure and a small piece of peptide of the transverse bonds. Based on computer molecular models, lysozyme has been proposed to distort the shape of one of the sugar rings in the chain, making it easier to break (other studies have suggested that different effects, such as electrostatic ones, are more important).
This molecule shows the possible structure of this distorted (green) ring which is bound by ester bond to a glutamic acid, at the top, of the active site of the lysozyme.
Normally, the sugar rings adopt a chair shape (zigzag) like the pink ring on the right.
Compare this ring with the one on the left, green, which has a shape folded up in the front, but flattened in the rest of the ring. This less stable structure is called half-chair.
In the following image the two rings are shown in the foreground to better appreciate their three-dimensional structure with half chair (green) and chair (pink).
148l: R. Kuroki, L. H. Weaver & B. W. Matthews (1993) A covalent enzyme-substrate intermediate with saccharide distortion in a mutant T4 lysozyme. Science 262, 2030-2033.
1l35: P. E. Pjura, M. Matsumura, J. A. Wozniak & B. W. Matthews (1990) Structure of a thermostable disulfide-bridge mutant of phage T4 lysozyme shows that an engineered cross-link in a flexible region does not increas the rigidity of the folded protein. Biochemistry 29, 2592-2598.
1lyd: D. R. Rose, J. Phipps, J. Michniewicz, G. I. Birnbaum, F. R. Ahmed, A. Muir, W. F. Anderson & S. Narang (1988) Crystal structure of T4-lysozyme generated from synthetic coding DNA in Escherichia coli. Protein Engineering 2, 277-282.
2lyz: R. Diamond (1974) Real-space refinement of the structure hen egg-while lysozyme. Journal of Molecular Biology 82, 371-391.