Anhydrosialidase

 

Intramolecular trans-sialidase (IT-sialidase) specifically cleaves the terminal N-acetyl-neuraminic acid-alpha-2-3-Galactose linkage in sialoglyco-conjugates. The released product is 2,7-anhydro-N-acetyl-neuraminic acid, indicating that the enzyme carries out an intramolecular reaction mechanism. This is in contrast to the hydrolytic sialidases whose product is free N-acetyl-neuraminic acid.

 

Reference Protein and Structure

Sequence
Q27701 UniProt (4.2.2.15) IPR026856 (Sequence Homologues) (PDB Homologues)
Biological species
Macrobdella decora (North American leech) Uniprot
PDB
1sll - SIALIDASE L FROM LEECH MACROBDELLA DECORA (2.0 Å) PDBe PDBsum 1sll
Catalytic CATH Domains
2.120.10.10 CATHdb (see all for 1sll)
Click To Show Structure

Enzyme Reaction (EC:4.2.2.15)

alpha-Neup5Ac-(2->8)-alpha-Neup5Ac-(2->8)-alpha-Neup5Ac
CHEBI:72328ChEBI
→
N-acetyl-2,7-anhydro-alpha-neuraminic acid
CHEBI:28695ChEBI
+
alpha-Neup5Ac-(2->8)-alpha-Neup5Ac
CHEBI:62097ChEBI
Alternative enzyme names: Anhydroneuraminidase, Sialglycoconjugate N-acylneuraminylhydrolase (2,7-cyclizing), Sialidase L,

Enzyme Mechanism

Introduction

In the first step, O2 of the substrate acquires a proton and leaves. Asp 318 either provides this proton directly or stabilises a proton from the solvent in the vicinity. The resulting positively charged oxocarbonium intermediate is stabilised by the negative side chains of Asp 318 and Glu 595, and also by a lone pair from the phenolic OH of Tyr 713. In the second step, the C7 hydroxyl attacks the oxocarbonium ion at C2. Asp 318 acts as a general base to deprotonate the attacking hydroxyl group.

Catalytic Residues Roles

UniProt PDB* (1sll)
Asp318 Asp318(238)A May protonate departing O2 of substrate, or at least stabilise a proton in the vicinity. Provides electrostatic stabilisation for the positively charged oxocarbonium ion. Accepts proton from the O7 hydroxyl group that attacks the oxocarbonium ion. proton acceptor, electrostatic stabiliser, proton donor
Glu595 Glu595(515)A Provides electrostatic stabilisation for the positively charged oxocarbonium ion. Also interacts with Tyr 713 to direct its O lone pair towards the substrate C2. electrostatic stabiliser
Tyr713 Tyr713(633)A Stabilises the oxocarbonium ion by directing a lone pair on the phenolic oxygen towards C2 of the substrate. electrostatic stabiliser
*PDB label guide - RESx(y)B(C) - RES: Residue Name; x: Residue ID in PDB file; y: Residue ID in PDB sequence if different from PDB file; B: PDB Chain; C: Biological Assembly Chain if different from PDB. If label is "Not Found" it means this residue is not found in the reference PDB.

Chemical Components

proton transfer, overall reactant used, overall product formed, heterolysis, intramolecular nucleophilic addition, native state of enzyme regenerated

References

  1. Luo Y et al. (1999), J Mol Biol, 285, 323-332. The 1.8 å structures of leech intramolecular trans-sialidase complexes: evidence of its enzymatic mechanism. DOI:10.1006/jmbi.1998.2345. PMID:9878409.
  2. Cremona ML et al. (1995), Gene, 160, 123-128. A single tyrosine differentiates active and inactive Trypanosoma cruzi trans-sialidases. DOI:10.1016/0378-1119(95)00175-6. PMID:7628705.

Step 1. Asp 318 protonates O2 of the substrate this causes the C2-O2 bond to be broken. The resulting oxocarbonium intermediate is stabilized by Asp 318, Glu595 and Tyr713.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Glu595(515)A electrostatic stabiliser
Tyr713(633)A electrostatic stabiliser
Asp318(238)A electrostatic stabiliser, proton donor

Chemical Components

proton transfer, overall reactant used, overall product formed, heterolysis

Step 2. The C7 hydroxyl attacks the oxocarbonium ion at C2, Asp 318 acts as a general base deprotonating the hydroxyl. Leading to the second product being formed.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Asp318(238)A electrostatic stabiliser
Glu595(515)A electrostatic stabiliser
Tyr713(633)A electrostatic stabiliser
Asp318(238)A proton acceptor

Chemical Components

proton transfer, ingold: intramolecular nucleophilic addition, overall product formed, native state of enzyme regenerated
Download: Image, Marvin File

Step 1. Asp 318 protonates O2 of the substrate this causes the C2-O2 bond to be broken. The resulting oxocarbonium intermediate is stabilized by Asp 318, Glu595 and Tyr713.

Step 2. The C7 hydroxyl attacks the oxocarbonium ion at C2, Asp 318 acts as a general base deprotonating the hydroxyl. Leading to the second product being formed.

Products.

Contributors

Steven Smith, Gemma L. Holliday, James Willey