PDBsum entry: 3edk

Hydrolase

PDB id: 3edk

Contents

Protein chains

597 a.a. *

Ligands

CE8 ×2

GOL ×4

Metals

_CA ×4

Waters ×1105

* Residue conservation analysis

PDB id:

3edk

Name:

Hydrolase

Title:

Structural base for cyclodextrin hydrolysis

Structure:

Cyclomaltodextrinase. Chain: a, b. Synonym: fspcmd. Engineered: yes. Mutation: yes

Source:

Flavobacterium sp. 92. Organism_taxid: 197856. Gene: cdase. Expressed in: escherichia coli. Expression_system_taxid: 562.

Resolution:

1.77Å R-factor: 0.176 R-free: 0.208

Authors:

S.Buedenbender,G.E.Schulz

Key ref:

S.Buedenbender and G.E.Schulz (2009). Structural base for enzymatic cyclodextrin hydrolysis. J Mol Biol, 385, 606-617. PubMed id: 19014948 DOI: 10.1016/j.jmb.2008.10.085

Date:

03-Sep-08 Release date: 03-Mar-09

Protein chains

Q8KKG0  (Q8KKG0_9FLAO) -  Cyclomaltodextrinase (Precursor)

Seq: 619 a.a.

Struc: 597 a.a.*

* PDB and UniProt seqs differ at 2 residue positions (black crosses)

Enzyme reactions

• Enzyme class: E.C.3.2.1.54 - Cyclomaltodextrinase.
• Reaction: Cyclomaltodextrin + H₂O = linear maltodextrin

Cyclomaltodextrin (Bound ligand (Het Group name = CE8) matches with 50.00% similarity) + H(2)O = linear maltodextrin

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

 Gene Ontology (GO) functional annotation 

• Biological process: metabolic process (2 terms)
• Biochemical function: catalytic activity (6 terms)

Added reference

DOI no: 10.1016/j.jmb.2008.10.085

J Mol Biol 385:606-617 (2009)

PubMed id: 19014948

Structural base for enzymatic cyclodextrin hydrolysis.

S.Buedenbender, G.E.Schulz.

ABSTRACT

Cyclodextrins resist hydrolysis by burying all bridge oxygens at their interior. Still, the rings can be opened by a small group of specialized enzymes, the cyclomaltodextrinases. Among them, the enzyme from Flavobacterium sp. no. 92 was mutated, crystallized and soaked with cyclodextrins, giving rise to four complex structures. One of them showed an alpha-cyclodextrin at the outer rim of the active center pocket. In the other complexes, alpha-, beta-and gamma-cyclodextrins were bound in a competent mode in the active center. The structures suggest that Arg464 functions as a chaperone guiding the substrates from the solvent into the active center. Over the last part of this pathway, the cyclodextrins bump on Phe274, which rotates the glucosyl group at subsite (+1) by about 120 degrees and fixes it in the new conformation. This induced fit was observed with all three major cyclodextrins. It makes the bridging oxygen between subsites (+1) and (-1) available for protonation by Glu340, which starts the hydrolysis. The mechanism resembles a spring-lock. The structural data were supplemented by activity measurements, quantifying the initial ring opening reaction for the major cyclodextrins and the transglucosylation activity for maltotetraose. Further activity data were collected for mutants splitting the tetrameric enzyme into dimers and for active center mutants.

Selected figure(s)

Figure 2.
Fig. 2. Stereoview of tetrameric FspCMD in crystal form I represented by a Cα trace. The horizontal twofold axis in the paper plane is crystallographic. The axes of the (βα)[8]-barrels are shown in red. The α-CD bound to one of the two independent subunits (15 min soak, Table 2) is given as a ball-and-stick model. The domains of the lower right-hand subunit are labeled. The view emphasizes the hollow structure of the tetramer and the small 530 Å^2 interface, which was dissociated by the mutation T49P (pink ball). Residue 237 of the 60 Å^2 B-domain moiety of the 530 Å^2 interface is marked (black ball); it is rather close to Phe274 at the active center. The large 1620 Å^2 interface connects the A, B and C domains. It has been disturbed by mutations at Ser399, His521 and Gly523 (yellow balls).

Figure 5.
Fig. 5. Stereoviews of cyclodextrins bound to inactive FspCMD mutants. Residues are shown with black Cα atoms. All 6-hydroxyls are emphasized by using large pink balls. (a) Comparison between two α-CD soaks. In crystal form I, the short 15 min soak leaves α-CD in a symmetrical conformation (yellow) at a general sugar-binding site 5 Å distant from the active center. The long 900 min soak in crystal form II deforms α-CD (blue), in particular at subsite (+1) (magenta), so that Glu340 (here Gln) can protonate the bridge oxygen (red dots). (b) The competent binding structures of α-CD (blue), β-CD (green) and γ-CD (purple) in crystal form II (chain-B). Note that the green Phe274 is visually obscured by the blue Phe274. The residue conformations of the 15 min α-CD soak are shown (yellow) but the respective α-CD molecule is deleted for clarity. The residue conformations of the unligated active enzyme are given in orange.

The above figures are reprinted by permission from Elsevier: J Mol Biol (2009, 385, 606-617) copyright 2009.

Figures were selected by the author.