PDBsum entry 1i2r

Transferase

PDB id: 1i2r

Contents

Protein chains

316 a.a. *

Waters ×313

* Residue conservation analysis

PDB id:

1i2r

Name:

Transferase

Title:

Crystal structure of escherichia coli transaldolase b mutant s176a

Structure:

Transaldolase b. Chain: a, b. Engineered: yes. Mutation: yes

Source:

Escherichia coli. Organism_taxid: 562. Gene: talb. Expressed in: escherichia coli. Expression_system_taxid: 562.

Biol. unit:

Dimer (from PQS)

Resolution:

2.10Å R-factor: 0.183 R-free: 0.203

Authors:

S.Thorell,J.Jia,G.Schneider

Key ref:

U.Schörken et al. (2001). Identification of catalytically important residues in the active site of Escherichia coli transaldolase. Eur J Biochem, 268, 2408-2415. PubMed id: 11298760 DOI: 10.1046/j.1432-1327.2001.02128.x

Date:

12-Feb-01 Release date: 09-May-01

Protein chains

P0A870  (TALB_ECOLI) -  Transaldolase B from Escherichia coli (strain K12)

Seq: 317 a.a.

Struc: 316 a.a.*

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

Enzyme reactions

• Enzyme class: E.C.2.2.1.2 - transaldolase.
• Reaction: D-sedoheptulose 7-phosphate + D-glyceraldehyde 3-phosphate = D-erythrose 4-phosphate + beta-D-fructose 6-phosphate

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

reference

DOI no: 10.1046/j.1432-1327.2001.02128.x

Eur J Biochem 268:2408-2415 (2001)

PubMed id: 11298760

Identification of catalytically important residues in the active site of Escherichia coli transaldolase.

U.Schörken, S.Thorell, M.Schürmann, J.Jia, G.A.Sprenger, G.Schneider.

ABSTRACT

The roles of invariant residues at the active site of transaldolase B from Escherichia coli have been probed by site-directed mutagenesis. The mutant enzymes D17A, N35A, E96A, T156A, and S176A were purified from a talB-deficient host and analyzed with respect to their 3D structure and kinetic behavior. X-ray analysis showed that side chain replacement did not induce unanticipated structural changes in the mutant enzymes. Three mutations, N35A, E96A, and T156A resulted mainly in an effect on apparent kcat, with little changes in apparent Km values for the substrates. Residues N35 and T156 are involved in the positioning of a catalytic water molecule at the active site and the side chain of E96 participates in concert with this water molecule in proton transfer during catalysis. Substitution of Ser176 by alanine resulted in a mutant enzyme with 2.5% residual activity. The apparent Km value for the donor substrate, fructose 6-phosphate, was increased nearly fivefold while the apparent Km value for the acceptor substrate, erythrose 4-phosphate remained unchanged, consistent with a function for S176 in the binding of the C1 hydroxyl group of the donor substrate. The mutant D17A showed a 300-fold decrease in kcat, and a fivefold increase in the apparent Km value for the acceptor substrate erythrose 4-phosphate, suggesting a role of this residue in carbon-carbon bond cleavage and stabilization of the carbanion/enamine intermediate.

Selected figure(s)

Figure 2.
Fig. 2. Stereo views of the final 2|Fo|-|Fc| electron density maps, contoured at 1 , of the transaldolase mutants D17A (A) and S176A (B).

Figure 4.
Fig. 4. Proposed reaction mechanism of transaldolase. The steps leading to the central carbanion/enamine intermediate are shown. The second half of the reaction, the addition of the acceptor substrate is in principle the reverse of the first half of the catalytic cycle and is therefore not included in the figure. For sake of clarity, only conserved amino-acid side chains proposed to participate in proton transfer during the reaction are shown.

The above figures are reprinted by permission from the Federation of European Biochemical Societies: Eur J Biochem (2001, 268, 2408-2415) copyright 2001.

Figures were selected by an automated process.