Lysozyme (glycosyl hydrolase 25 family)
Cpl-1 lysin from Bacteriophage cp-1 catalyses the hydrolysis of the 1,4-beta-linkages between N-acetyl-D-glucosamine and N-acetylmuramic acid in peptidoglycan heteropolymers of prokaryote cell walls. On binding to choline, the enzymes geometry changes, its specificity increases, and catalytic activity is increased. Cpl-1 is used against bacteria such as Streptococcus pneumoniae (which causes pneumonia), as it kills the bacteria by breaking down its cell wall.
Reference Protein and Structure
- Sequence
-
P15057
(3.2.1.17)
(Sequence Homologues)
(PDB Homologues)
- Biological species
-
Streptococcus phage Cp-1 (Virus)
- PDB
-
1oba
- Multimodular Pneumococcal Cell Wall Endolysin from phage Cp-1 complexed with choline
(2.45 Å)
- Catalytic CATH Domains
-
3.20.20.80
(see all for 1oba)
Enzyme Reaction (EC:3.2.1.17)
Enzyme Mechanism
Introduction
More recent evidence suggests a mechanism which proceeds via an oxazoline ion. Glu94 donates a proton to the glycosidic oxygen causing the cleavage of the glycosidic bond and the formation of the oxazoline ion. A water molecule is then activated by Glu94 which hydrolyzes the oxazoline ion.
Catalytic Residues Roles
| UniProt | PDB* (1oba) | ||
| Asp10 | Asp10A | Stabilizes the oxazoline ion | electrostatic stabiliser |
| Glu94 | Glu94A | Protonates the leaving group and activates the water. | proton acceptor, proton donor, activator, increase nucleophilicity, promote heterolysis |
Chemical Components
overall product formed, overall reactant used, cyclisation, proton transfer, intramolecular nucleophilic substitution, decyclisation, hydrolysis, bimolecular nucleophilic substitution, native state of enzyme regeneratedReferences
- Martinez-Fleites C et al. (2009), Carbohydr Res, 344, 1753-1757. The crystal structure of a family GH25 lysozyme from Bacillus anthracis implies a neighboring-group catalytic mechanism with retention of anomeric configuration. DOI:10.1016/j.carres.2009.06.001. PMID:19595298.
Step 1. Glu94 donates a proton to the glycosidic oxygen. The glycosidic bond is cleaved when the oxygen of the N-acetyl group attacks C1. The subsequent oxazoline ion formed is stabilized by Asp10.
Download: Image, Marvin FileCatalytic Residues Roles
| Residue | Roles |
|---|---|
| Asp10A | electrostatic stabiliser |
| Glu94A | promote heterolysis, proton donor |
Chemical Components
overall product formed, overall reactant used, cyclisation, proton transfer, ingold: intramolecular nucleophilic substitution
Step 2. Glu94 activates a water molecule which attacks C1 and the oxazoline ion collapses into the product.
Download: Image, Marvin FileCatalytic Residues Roles
| Residue | Roles |
|---|---|
| Glu94A | activator, increase nucleophilicity |
| Asp10A | electrostatic stabiliser |
| Glu94A | proton acceptor |
Chemical Components
decyclisation, overall product formed, proton transfer, hydrolysis, ingold: bimolecular nucleophilic substitution, native state of enzyme regenerated
Download: Introduction
Asp 10 acts as a general base, activating a water molecule for nucleophilic attack at the carbon atom of the glycosidic bond. As the glycosidic bond cleaves, protonated Glu 94 acts as a general acid by protonating the leaving group oxygen atom.
Catalytic Residues Roles
| UniProt | PDB* (1oba) | ||
| Asp10 | Asp10A | Asp 10 acts as a general base by deprotonating a water molecule, activating it for nucleophilic attack on the carbon atom of the glycosidic bond. | proton acceptor, proton donor, activator, electrostatic stabiliser, increase nucleophilicity |
| Asp92 | Asp92A | Asp 92 allows protons to move freely between itself and Glu 94, thus ensuring regeneration of the protonated state of Glu 94. | proton donor |
| Glu94 | Glu94A | The protonated form of Glu 94 acts as a general acid, by protonating the leaving group oxygen atom as the glycosidic bond is broken. | promote heterolysis, proton acceptor, proton donor |
| Asp182 | Asp182A | Low barrier hydrogen bonds allow the free transfer of protons between Asp 182 and Asp 10, ensuring the regeneration of the deprotonated state of Asp 10. | proton acceptor |
Chemical Components
overall product formed, overall reactant used, proton transfer, heterolysis, bimolecular nucleophilic addition, native state of enzyme regeneratedReferences
- Hermoso JA et al. (2003), Structure, 11, 1239-1249. Structural Basis for Selective Recognition of Pneumococcal Cell Wall by Modular Endolysin from Phage Cp-1. DOI:10.1016/j.str.2003.09.005. PMID:14527392.
Step 1. Glu94 donates a proton to the glycosidic oxygen which promotes the cleavage of the glycosidic bond.
Download: Image, Marvin FileCatalytic Residues Roles
| Residue | Roles |
|---|---|
| Asp10A | electrostatic stabiliser |
| Glu94A | promote heterolysis, proton donor |
Chemical Components
overall product formed, overall reactant used, proton transfer, heterolysis
Step 2. Asp10 activates a water molecule which attacks the oxycarbenium ion.
Download: Image, Marvin FileCatalytic Residues Roles
| Residue | Roles |
|---|---|
| Asp10A | activator, increase nucleophilicity, proton acceptor |
Chemical Components
overall product formed, proton transfer, ingold: bimolecular nucleophilic addition
Step 3. Asp92 and Asp182 regenerate the catalytic residues in their native states.
Download: Image, Marvin FileCatalytic Residues Roles
| Residue | Roles |
|---|---|
| Glu94A | proton acceptor |
| Asp182A | proton acceptor |
| Asp10A | proton donor |
| Asp92A | proton donor |
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