Glucan 1,4-alpha-maltohydrolase

 

Alpha-amylases, which belong to glycoside hydrolase family 13, are 1,4-alpha-D-glucan glucanohydrolases, which degrade both the branched and unbranched forms of starch by cleaving the internal alpha-1,4 bonds connecting the glucose monomers. The products of these reactions are maltose and maltotriose, which are further degraded to glucose by maltases. One atom of calcium is required to bind to each protein molecule to allow it to function, but excess calcium can inhibit activity by binding to amino acids that are required for the catalytic activity of the enzyme.

 

Reference Protein and Structure

Sequence
P19531 UniProt (3.2.1.133) IPR006046 (Sequence Homologues) (PDB Homologues)
Biological species
Geobacillus stearothermophilus (Bacteria) Uniprot
PDB
1qho - FIVE-DOMAIN ALPHA-AMYLASE FROM BACILLUS STEAROTHERMOPHILUS, MALTOSE/ACARBOSE COMPLEX (1.7 Å) PDBe PDBsum 1qho
Catalytic CATH Domains
3.20.20.80 CATHdb (see all for 1qho)
Cofactors
Calcium(2+) (1)
Click To Show Structure

Enzyme Reaction (EC:3.2.1.133)

water
CHEBI:15377ChEBI
+
alpha-maltotetraose
CHEBI:61988ChEBI
→
alpha-maltose
CHEBI:18167ChEBI
+
alpha-maltose
CHEBI:18167ChEBI
Alternative enzyme names: Maltogenic alpha-amylase, 1,4-alpha-D-glucan alpha-maltohydrolase,

Enzyme Mechanism

Introduction

This enzyme catalyses glycoside hydrolysis with net retention of anomeric configuration by a double-displacement mechanism. In this mechanism, Asp228 initiates a nucleophilic attack on the anomeric carbon, displacing one of the products with concomitant deprotonation of Glu256. Glu256 then abstracts a proton from the catalytic water molecule, which in turn attacks the anomeric carbon, displacing Asp228.

Catalytic Residues Roles

UniProt PDB* (1qho)
Asp261 Asp228A Acts as the catalytic nucleophile. covalent catalysis
Glu289 Glu256A Acts as a general acid/base. This residues works as the general acid catalyst that donates a proton to O4 (glucosidic bond oxygen) in the first step of the general acid–base catalysis. It then abstracts the proton from the ncucleophilic water molecule. proton shuttle (general acid/base)
Asp362 Asp329A Helps stabilise the transition states and reactive intermediates formed during the course of the reaction. The residue helps bind the substrate is a twisted and deformed conformation of the glucose ring at position -1 (Glc4), which enhances the reactivity. transition state 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

References

  1. Hasegawa K et al. (1999), Protein Eng, 12, 819-824. Roles of catalytic residues in  -amylases as evidenced by the structures of the product-complexed mutants of a maltotetraose-forming amylase. DOI:10.1093/protein/12.10.819. PMID:10556241.
  2. van der Maarel MJ et al. (2002), J Biotechnol, 94, 137-155. Properties and applications of starch-converting enzymes of the alpha-amylase family. PMID:11796168.
  3. Qian M et al. (2001), Biochemistry, 40, 7700-7709. Enzyme-Catalyzed Condensation Reaction in a Mammalian α-Amylase. High-Resolution Structural Analysis of an Enzyme−Inhibitor Complex†. DOI:10.1021/bi0102050.
  4. Mezaki Y et al. (2001), Biosci Biotechnol Biochem, 65, 222-225. Crystallization and Structural Analysis of Intact Maltotetraose-forming Exo-amylase from Pseudomonas stutzeri. DOI:10.1271/bbb.65.222. PMID:11272837.
  5. van der Veen BA et al. (2000), Biochim Biophys Acta, 1543, 336-360. Engineering of cyclodextrin glycosyltransferase reaction and product specificity. PMID:11150613.
  6. Dauter Z et al. (1999), Biochemistry, 38, 8385-8392. X-ray structure of Novamyl, the five-domain "maltogenic" alpha-amylase from Bacillus stearothermophilus: maltose and acarbose complexes at 1.7A resolution. DOI:10.1021/bi990256l. PMID:10387084.
  7. Morishita Y et al. (1997), J Mol Biol, 267, 661-672. Crystal structure of a maltotetraose-formingexo-amylase from Pseudomonas stutzeri. DOI:10.1006/jmbi.1996.0887. PMID:9126844.
  8. Yoshioka Y et al. (1997), J Mol Biol, 271, 619-628. Crystal structures of a mutant maltotetraose-forming exo-amylase cocrystallized with maltopentaose 1 1Edited by R. Huber. DOI:10.1006/jmbi.1997.1222. PMID:9281429.

Step 1.

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Catalytic Residues Roles

Residue Roles
Glu256A proton shuttle (general acid/base)
Asp228A covalent catalysis
Asp329A transition state stabiliser

Chemical Components

Step 1.

Contributors

Alex Gutteridge, Craig Porter, Gemma L. Holliday