PDBsum entry 4zu7

Isomerase

PDB id: 4zu7

Contents

Protein chains

(+ 2 more) 137 a.a.

Ligands

TYD ×8

PXN

Waters ×179

PDB id:

4zu7

Name:

Isomerase

Title:

X-ray structure if the qdta 3,4-ketoisomerase from thermoanaerobacterium thermosaccharolyticum, double mutant y17r/r97h, in complex with tdp

Structure:

Qdta. Chain: a, b, c, d, e, f, g, h. Engineered: yes. Mutation: yes

Source:

Thermoanaerobacterium thermosaccharolyticum. Organism_taxid: 1517. Gene: qdta. Expressed in: escherichia coli. Expression_system_taxid: 469008.

Resolution:

2.30Å R-factor: 0.196 R-free: 0.256

Authors:

J.B.Thoden,E.Vinogradov,M.Gilbert,A.J.Salinger,H.M.Holden

Key ref:

J.B.Thoden et al. (2015). Bacterial Sugar 3,4-Ketoisomerases: Structural Insight into Product Stereochemistry. Biochemistry, 54, 4495-4506. PubMed id: 26125548 DOI: 10.1021/acs.biochem.5b00541

Date:

15-May-15 Release date: 15-Jul-15

Protein chains

Q6TFC5  (Q6TFC5_THETR) -  QdtA from Thermoanaerobacterium thermosaccharolyticum

Seq: 136 a.a.

Struc: 137 a.a.*

* PDB and UniProt seqs differ at 6 residue positions (black crosses)

DOI no: 10.1021/acs.biochem.5b00541

Biochemistry 54:4495-4506 (2015)

PubMed id: 26125548

Bacterial Sugar 3,4-Ketoisomerases: Structural Insight into Product Stereochemistry.

J.B.Thoden, E.Vinogradov, M.Gilbert, A.J.Salinger, H.M.Holden.

ABSTRACT

3-Acetamido-3,6-dideoxy-d-galactose (Fuc3NAc) and 3-acetamido-3,6-dideoxy-d-glucose (Qui3NAc) are unusual sugars found on the lipopolysaccharides of Gram-negative bacteria and on the S-layers of Gram-positive bacteria. The 3,4-ketoisomerases, referred to as FdtA and QdtA, catalyze the third steps in the respective biosynthetic pathways for these sugars. Whereas both enzymes utilize the same substrate, the stereochemistries of their products are different. Specifically, the hydroxyl groups at the hexose C-4' positions assume the "galactose" and "glucose" configurations in the FdtA and QdtA products, respectively. In 2007 we reported the structure of the apoform of FdtA from Aneurinibacillus thermoaerophilus, which was followed in 2014 by the X-ray analysis of QdtA from Thermoanaerobacterium thermosaccharolyticum as a binary complex. Both of these enzymes belong to the cupin superfamily. Here we report a combined structural and enzymological study to explore the manner in which these enzymes control the stereochemistry of their products. Various site-directed mutant proteins of each enzyme were constructed, and their dTDP-sugar products were analyzed by NMR spectroscopy. In addition, the kinetic parameters for these protein variants were measured, and the structure of one, namely, the QdtA Y17R/R97H double mutant form, was determined to 2.3-Å resolution. Finally, in an attempt to obtain a model of FdtA with a bound dTDP-linked sugar, the 3,4-ketoisomerase domain of a bifunctional enzyme from Shewanella denitrificans was cloned, purified, and crystallized in the presence of a dTDP-linked sugar analogue. Taken together, the results from this investigation demonstrate that it is possible to convert a "galacto" enzyme into a "gluco" enzyme and vice versa.