PDBsum entry 1v3l

Transferase

PDB id: 1v3l

Contents

Protein chains

686 a.a. *

Ligands

GAL-GLD

GLC-GLD

GLC-ACI ×2

Metals

_CA ×4

Waters ×504

* Residue conservation analysis

PDB id:

1v3l

Name:

Transferase

Title:

Crystal structure of f283l mutant cyclodextrin glycosyltransferase complexed with a pseudo-tetraose derived from acarbose

Structure:

Cyclomaltodextrin glucanotransferase. Chain: a, b. Synonym: cyclodextrin-glycosyltransferase, cgtase. Engineered: yes. Mutation: yes

Source:

Bacillus sp.. Organism_taxid: 1410. Strain: 1011. Expressed in: escherichia coli. Expression_system_taxid: 562.

Resolution:

2.10Å R-factor: 0.169 R-free: 0.232

Authors:

R.Kanai,K.Haga,T.Akiba,K.Yamane,K.Harata

Key ref:

R.Kanai et al. (2004). Role of Phe283 in enzymatic reaction of cyclodextrin glycosyltransferase from alkalophilic Bacillus sp.1011: Substrate binding and arrangement of the catalytic site. Protein Sci, 13, 457-465. PubMed id: 14739329 DOI: 10.1110/ps.03408504

Date:

03-Nov-03 Release date: 03-Aug-04

Protein chains

P05618  (CDGT_BACS0) -  Cyclomaltodextrin glucanotransferase from Bacillus sp. (strain 1011)

Seq: 713 a.a.

Struc: 686 a.a.*

* PDB and UniProt seqs differ at 3 residue positions (black crosses)

Enzyme reactions

• Enzyme class: E.C.2.4.1.19 - cyclomaltodextrin glucanotransferase.
• Reaction: Degrades starch to cyclodextrins by formation of a 1,4-alpha-D- glucosidic bond.

DOI no: 10.1110/ps.03408504

Protein Sci 13:457-465 (2004)

PubMed id: 14739329

Role of Phe283 in enzymatic reaction of cyclodextrin glycosyltransferase from alkalophilic Bacillus sp.1011: Substrate binding and arrangement of the catalytic site.

R.Kanai, K.Haga, T.Akiba, K.Yamane, K.Harata.

ABSTRACT

Cyclodextrin glycosyltransferase (CGTase) belonging to the alpha-amylase family mainly catalyzes transglycosylation and produces cyclodextrins from starch and related alpha-1,4-glucans. The catalytic site of CGTase specifically conserves four aromatic residues, Phe183, Tyr195, Phe259, and Phe283, which are not found in alpha-amylase. To elucidate the structural role of Phe283, we determined the crystal structures of native and acarbose-complexed mutant CGTases in which Phe283 was replaced with leucine (F283L) or tyrosine (F283Y). The temperature factors of the region 259-269 in native F283L increased >10 A(2) compared with the wild type. The complex formation with acarbose not only increased the temperature factors (>10 A(2)) but also changed the structure of the region 257-267. This region is stabilized by interactions of Phe283 with Phe259 and Leu260 and plays an important role in the cyclodextrin binding. The conformation of the side-chains of Glu257, Phe259, His327, and Asp328 in the catalytic site was altered by the mutation of Phe283 with leucine, and this indicates that Phe283 partly arranges the structure of the catalytic site through contacts with Glu257 and Phe259. The replacement of Phe283 with tyrosine decreased the enzymatic activity in the basic pH range. The hydroxyl group of Tyr283 forms hydrogen bonds with the carboxyl group of Glu257, and the pK(a) of Glu257 in F283Y may be lower than that in the wild type.

Selected figure(s)

Figure 1.
Figure 1. Absolute starch degrading activity. pH Profiles for F283L (Nakamura et al. 1994 [solid circles]), F283Y mutant (solid squares), and wild-type CGTase (solid diamonds).

Figure 4.
Figure 4. Comparison of structures and temperature factors. (A) Difference of isotropic temperature factors for equivalent C atoms between F283L and wild-type CGTase. (B) Difference of isotropic temperature factors for equivalent C atoms between F283L_ACA and F283L. (C) Positional difference for equivalent C atoms in domain A (residues 1-138 and 204-406) between F283L_ACA and F283L.

The above figures are reprinted by permission from the Protein Society: Protein Sci (2004, 13, 457-465) copyright 2004.

Figures were selected by the author.