User:Jsolomon580/sandbox

Function & Mechanism
The enzyme functions by attacking peptidoglycans (found in the cell walls of bacteria, especially Gram-positive bacteria) and hydrolyzing the glycosidic bond that connects N-acetylmuramic acid with the fourth carbon atom of N-acetylglucosamine. It does this by binding to the peptidoglycan molecule in the binding site within the prominent cleft between its two domains. This causes the substrate molecule to adopt a strained conformation similar to that of the transition state. According to Phillips-Mechanism, the lysozyme binds to a hexasaccharide. The lysozyme then distorts the fourth sugar in hexasaccharide (the D ring) into a half-chair conformation. In this stressed state, the glycosidic bond is easily broken. The amino acid side-chains glutamic acid 35 (Glu35) and aspartate 52 (Asp52) have been found to be critical to the activity of this enzyme. Glu35 acts as a proton donor to the glycosidic bond, cleaving the C-O bond in the substrate, whereas Asp52 acts as a nucleophile to generate a glycosyl enzyme intermediate. The glycosyl enzyme intermediate then reacts with a water molecule, to give the product of hydrolysis and leaving the enzyme unchanged.

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 Koshland mechanism is ~30 kcal/mol more stable than the ionic intermediate from the Phillips mechanism. These calculation 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.





3. Bowman A.L.,Grant, I. M. and Mulholland, A. J. (2008). "QM/MM simulations predict a covalent intermediate in the hen egg white [null lysozyme] reaction with its natural substrate" ''Chem. Commun''., 37, 4425. DOI: 10.1039/B810099C

4. Vocaldo, D.J., Davies, G.J., Laine, R. and Withers, S. G.(2001) "Catalysis by hen egg-white lysozyme Proceeds via a covalent intermediate" Nature. 412, 835.