PDBsum entry 2dw7

Lyase

PDB id: 2dw7

Contents

Protein chains

(+ 10 more) 388 a.a. *

Ligands

SRT ×16

Metals

_MG ×16

Waters ×449

* Residue conservation analysis

PDB id:

2dw7

Name:

Lyase

Title:

Crystal structure of d-tartrate dehydratase from bradyrhizobium japonicum complexed with mg++ and meso-tartrate

Structure:

Bll6730 protein. Chain: a, b, c, d, e, f, g, h, i, j, k, l, m, n, o, p. Synonym: d-tartrate dehydratase. Engineered: yes

Source:

Bradyrhizobium japonicum. Organism_taxid: 375. Expressed in: escherichia coli bl21(de3). Expression_system_taxid: 469008.

Resolution:

2.50Å R-factor: 0.241 R-free: 0.265

Authors:

A.A.Fedorov,E.V.Fedorov,W.S.Yew,B.M.Wood,J.A.Gerlt,S.C.Almo

Key ref:

W.S.Yew et al. (2006). Evolution of enzymatic activities in the enolase superfamily: D-tartrate dehydratase from Bradyrhizobium japonicum. Biochemistry, 45, 14598-14608. PubMed id: 17144653 DOI: 10.1021/bi061688g

Date:

07-Aug-06 Release date: 19-Dec-06

Protein chains

Q89FH0  (TARD_BRADU) -  D(-)-tartrate dehydratase from Bradyrhizobium diazoefficiens (strain JCM 10833 / BCRC 13528 / IAM 13628 / NBRC 14792 / USDA 110)

Seq: 389 a.a.

Struc: 388 a.a.

Enzyme reactions

• Enzyme class: E.C.4.2.1.81 - D(-)-tartrate dehydratase.
• Reaction: (S,S)-tartrate = oxaloacetate + H2O

(S,S)-tartrate (Bound ligand (Het Group name = SRT) corresponds exactly) = oxaloacetate + H2O

• Cofactor: Fe cation

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

Added reference

DOI no: 10.1021/bi061688g

Biochemistry 45:14598-14608 (2006)

PubMed id: 17144653

Evolution of enzymatic activities in the enolase superfamily: D-tartrate dehydratase from Bradyrhizobium japonicum.

W.S.Yew, A.A.Fedorov, E.V.Fedorov, B.M.Wood, S.C.Almo, J.A.Gerlt.

ABSTRACT

We focus on the assignment of function to and elucidation of structure-function relationships for a member of the mechanistically diverse enolase superfamily encoded by the Bradyrhizobium japonicum genome (bll6730; GI:27381841). As suggested by sequence alignments, the active site contains the same functional groups found in the active site of mandelate racemase (MR) that catalyzes a 1,1-proton transfer reaction: two acid/base catalysts, Lys 184 at the end of the second beta-strand, and a His 322-Asp 292 dyad at the ends of the seventh and sixth beta-strands, respectively, as well as ligands for an essential Mg2+, Asp 213, Glu 239, and Glu 265 at the ends of the third, fourth, and fifth beta-strands, respectively. We screened a library of 46 acid sugars and discovered that only d-tartrate is dehydrated, yielding oxaloacetate as product. The kinetic constants (kcat = 7.3 s(-1); kcat/KM = 8.5 x 10(4) M(-1) s(-1)) are consistent with assignment of the d-tartrate dehydratase (TarD) function. The kinetic phenotypes of mutants as well as the structures of liganded complexes are consistent with a mechanism in which Lys 184 initiates the reaction by abstraction of the alpha-proton to generate a Mg2+-stabilized enediolate intermediate, and the vinylogous beta-elimination of the 3-OH group is general acid-catalyzed by the His 322, accomplishing the anti-elimination of water. The replacement of the leaving group by solvent-derived hydrogen is stereorandom, suggesting that the enol tautomer of oxaloacetate is the product; this expectation was confirmed by its observation by 1H NMR spectroscopy. Thus, the TarD-catalyzed reaction is a "simple" extension of the two-step reaction catalyzed by MR: base-catalyzed proton abstraction to generate a Mg2+-stabilized enediolate intermediate followed by acid-catalyzed decomposition of that intermediate to yield the product.