PDBsum entry: 2ber

Hydrolase

PDB id: 2ber

Contents

Protein chain

601 a.a. *

Ligands

SLB

Metals

_NA

Waters ×837

* Residue conservation analysis

PDB id:

2ber

Name:

Hydrolase

Title:

Y370g active site mutant of the sialidase from micromonospora viridifaciens in complex with beta-neu5ac (sialic acid).

Structure:

Bacterial sialidase. Chain: a. Fragment: residues 47-647. Synonym: neuraminidase. Engineered: yes. Mutation: yes

Source:

Micromonospora viridifaciens. Organism_taxid: 1881. Atcc: 31146. Expressed in: escherichia coli. Expression_system_taxid: 469008.

Resolution:

1.80Å R-factor: 0.143 R-free: 0.199

Authors:

S.Newstead,J.N.Watson,A.J.Bennet,G.L.Taylor

Key ref:

S.Newstead et al. (2005). Structure and mechanism of action of an inverting mutant sialidase. Biochemistry, 44, 9117-9122. PubMed id: 15966735 DOI: 10.1021/bi050517t

Date:

30-Nov-04 Release date: 04-Apr-05

Protein chain

Q02834  (NANH_MICVI) -  Sialidase

Seq: 647 a.a.

Struc: 601 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

Enzyme reactions

• Enzyme class: E.C.3.2.1.18 - Exo-alpha-sialidase.
• Reaction: Hydrolysis of alpha-(2->3)-, alpha-(2->6)-, alpha-(2->8)-glycosidic linkages of terminal sialic residues in oligosaccharides, glycoproteins, glycolipids, colominic acid and synthetic substrates.

 Gene Ontology (GO) functional annotation 

• Cellular component: extracellular region (1 term)
• Biological process: metabolic process (2 terms)
• Biochemical function: hydrolase activity (6 terms)

DOI no: 10.1021/bi050517t

Biochemistry 44:9117-9122 (2005)

PubMed id: 15966735

Structure and mechanism of action of an inverting mutant sialidase.

S.Newstead, J.N.Watson, T.L.Knoll, A.J.Bennet, G.Taylor.

ABSTRACT

Mutagenesis of the conserved tyrosine (Y370) of the Micromonospora viridifaciens sialidase to small amino acids changes the mechanism of catalysis from retention of anomeric configuration to inversion [Watson, J. N., et al. (2003) Biochemistry 42, 12682-12690]. For the Y370G mutant enzyme-catalyzed hydrolysis of a series of aryl sialosides and 3'-sialyllactose, the derived Brønsted parameters (beta(lg)) on k(cat) and k(cat)/K(m) are -0.63 +/- 0.05 and -0.80 +/- 0.08, respectively. Thus, for the Y370G enzyme, glycosidic C-O bond cleavage is rate-determining. Analysis of the activity of the Y370G mutant and wild-type enzymes against a substrate [3,4-dihydro-2H-pyrano[3,2-c]pyridinium whose hydrolysis cannot be accelerated by acid catalysis is consistent with these reactions proceeding via S(N)1 and S(N)2 mechanisms, respectively. The overall structure of the Y370G mutant sialidase active site is very similar to the previously reported wild-type structure [Gaskell, A., et al. (1995) Structure 3, 1197-1205], although removal of the tyrosine residue creates two significant changes to the active site. First, the anomeric oxygen atom of the hydrolysis product (beta-N-acetylneuraminic acid) and four water molecules bind in the large cavity created by the Y370G mutation. Second, the side chain of Asn310 moves to make a strong hydrogen bond to one of the bound water molecules.