PDBsum entry 2ixi

Isomerase

PDB id: 2ixi

Contents

Protein chains

184 a.a. *

Ligands

TYD ×2

SRT

Waters ×455

* Residue conservation analysis

PDB id:

2ixi

Name:

Isomerase

Title:

Rmlc p aeruginosa with dtdp-xylose

Structure:

Dtdp-4-dehydrorhamnose 3,5-epimerase. Chain: a, b. Synonym: dtdp-4-keto-6-deoxyglucose 3,5-epimerase. Engineered: yes

Source:

Pseudomonas aeruginosa. Organism_taxid: 287. Expressed in: escherichia coli. Expression_system_taxid: 562

Biol. unit:

Dimer (from PDB file)

Resolution:

1.80Å R-factor: 0.188 R-free: 0.237

Authors:

C.Dong,J.H.Naismith

Key ref:

C.Dong et al. (2007). RmlC, a C3' and C5' carbohydrate epimerase, appears to operate via an intermediate with an unusual twist boat conformation. J Mol Biol, 365, 146-159. PubMed id: 17046787 DOI: 10.1016/j.jmb.2006.09.063

Date:

08-Jul-06 Release date: 11-Jul-06

Protein chains

Q9HU21  (RMLC_PSEAE) -  dTDP-4-dehydrorhamnose 3,5-epimerase from Pseudomonas aeruginosa (strain ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1)

Seq: 181 a.a.

Struc: 184 a.a.

Enzyme reactions

• Enzyme class: E.C.5.1.3.13 - dTDP-4-dehydrorhamnose 3,5-epimerase.
• Pathway: 6-Deoxyhexose Biosynthesis
• Reaction: dTDP-4-dehydro-6-deoxy-alpha-D-glucose = dTDP-4-dehydro-beta-L-rhamnose

dTDP-4-dehydro-6-deoxy-alpha-D-glucose (Bound ligand (Het Group name = TYD) matches with 71.43% similarity) = dTDP-4-dehydro-beta-L-rhamnose

• Cofactor: NAD(+)

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

Added reference

DOI no: 10.1016/j.jmb.2006.09.063

J Mol Biol 365:146-159 (2007)

PubMed id: 17046787

RmlC, a C3' and C5' carbohydrate epimerase, appears to operate via an intermediate with an unusual twist boat conformation.

C.Dong, L.L.Major, V.Srikannathasan, J.C.Errey, M.F.Giraud, J.S.Lam, M.Graninger, P.Messner, M.R.McNeil, R.A.Field, C.Whitfield, J.H.Naismith.

ABSTRACT

The striking feature of carbohydrates is their constitutional, conformational and configurational diversity. Biology has harnessed this diversity and manipulates carbohydrate residues in a variety of ways, one of which is epimerization. RmlC catalyzes the epimerization of the C3' and C5' positions of dTDP-6-deoxy-D-xylo-4-hexulose, forming dTDP-6-deoxy-L-lyxo-4-hexulose. RmlC is the third enzyme of the rhamnose pathway, and represents a validated anti-bacterial drug target. Although several structures of the enzyme have been reported, the mechanism and the nature of the intermediates have remained obscure. Despite its relatively small size (22 kDa), RmlC catalyzes four stereospecific proton transfers and the substrate undergoes a major conformational change during the course of the transformation. Here we report the structure of RmlC from several organisms in complex with product and product mimics. We have probed site-directed mutants by assay and by deuterium exchange. The combination of structural and biochemical data has allowed us to assign key residues and identify the conformation of the carbohydrate during turnover. Clear knowledge of the chemical structure of RmlC reaction intermediates may offer new opportunities for rational drug design.

Selected figure(s)

Figure 1.
Figure 1. (a) R = OdTDP. The RmlC reaction converts dTDP-6-deoxy-D-xylo-4-hexulose to dTDP-6-deoxy-L-lyxo-4-hexulose (thick arrow). This process involves a ring flip as well as epimerization. The steps are shown according to the current convention, however, this route goes through some very high energy intermediates (notably the C1′, C3′, C5′ triaxial product). (b) R = OGPD. The GME reaction converts GDP-D-mannose to GDP-L-galactose. Shown boxed is the predicted twist boat intermediate. The dominant order of the epimerization (C5′ first) and the ring flipped form of GDP-L-ribo-4-hexulose were determined experimentally.^24 (c) R = OdTDP. Substrate and product mimics employed in this study.

Figure 6.
Figure 6. (a) R = OdTDP. A possible mechanism for RmlC based on structural and biochemical data. The key active site residues are shown, the H65 is the catalytic base for both epimerizations, K73 stabilizes the enolate and Y134 acts as the acid for the first epimerization. The mono-epimerized intermediate is shown boxed and has the equatorial linkage between carbohydrate ring and nucleotide. It cannot proceed directly to product because the C3′ proton is only sufficiently acidic when it is orthogonal to the plane of the carbonyl function. (b) R = OdTDP. An alternative route for the first epimerization using a twist boat form of substrate, the mono epimerized intermediate is shown boxed. The apparent preference of RmlC for the equatorial linked sugar nucleotide suggests that this is a possibility.

The above figures are reprinted by permission from Elsevier: J Mol Biol (2007, 365, 146-159) copyright 2007.

Figures were selected by an automated process.