PDBsum entry 2uzh

Lyase

PDB id: 2uzh

Contents

Protein chains

153 a.a. *

Ligands

CDP ×3

EDO ×2

GOL ×3

IPE

PEG ×2

ACT

Metals

_ZN ×3

Waters ×272

* Residue conservation analysis

PDB id:

2uzh

Name:

Lyase

Title:

Mycobacterium smegmatis 2c-methyl-d-erythritol-2,4-cyclodiphosphate synthase (ispf)

Structure:

2c-methyl-d-erythritol 2,4-cyclodiphosphate synthase. Chain: a, b, c. Engineered: yes. Other_details: each contains a zn cation, which is required for activity, and cdp, a fragment of substrate.

Source:

Mycobacterium smegmatis. Organism_taxid: 1772. Strain: mc(2)155. Atcc: 700084. Expressed in: escherichia coli. Expression_system_taxid: 469008.

Resolution:

2.20Å R-factor: 0.165 R-free: 0.206

Authors:

L.Buetow,A.C.Brown,T.Parish,W.N.Hunter

Key ref:

L.Buetow et al. (2007). The structure of Mycobacteria 2C-methyl-D-erythritol-2,4-cyclodiphosphate synthase, an essential enzyme, provides a platform for drug discovery. Bmc Struct Biol, 7, 68. PubMed id: 17956607

Date:

27-Apr-07 Release date: 06-Nov-07

Protein chains

A0R559  (A0R559_MYCS2) -  2-C-methyl-D-erythritol 2,4-cyclodiphosphate synthase from Mycolicibacterium smegmatis (strain ATCC 700084 / mc(2)155)

Seq: 163 a.a.

Struc: 153 a.a.

Enzyme reactions

• Enzyme class: E.C.4.6.1.12 - 2-C-methyl-D-erythritol 2,4-cyclodiphosphate synthase.
• Reaction: 4-CDP-2-C-methyl-D-erythritol 2-phosphate = 2-C-methyl-D-erythritol 2,4- cyclic diphosphate + CMP

4-CDP-2-C-methyl-D-erythritol 2-phosphate = 2-C-methyl-D-erythritol 2,4- cyclic diphosphate + CMP (Bound ligand (Het Group name = CDP) matches with 84.00% similarity)

• Cofactor: Mn(2+) or Mg(2+)

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

Key reference

Bmc Struct Biol 7:68 (2007)

PubMed id: 17956607

The structure of Mycobacteria 2C-methyl-D-erythritol-2,4-cyclodiphosphate synthase, an essential enzyme, provides a platform for drug discovery.

L.Buetow, A.C.Brown, T.Parish, W.N.Hunter.

ABSTRACT

BACKGROUND: The prevalence of tuberculosis, the prolonged and expensive treatment that this disease requires and an increase in drug resistance indicate an urgent need for new treatments. The 1-deoxy-D-xylulose 5-phosphate pathway of isoprenoid precursor biosynthesis is an attractive chemotherapeutic target because it occurs in many pathogens, including Mycobacterium tuberculosis, and is absent from humans. To underpin future drug development it is important to assess which enzymes in this biosynthetic pathway are essential in the actual pathogens and to characterize them. RESULTS: The fifth enzyme of this pathway, encoded by ispF, is 2C-methyl-D-erythritol-2,4-cyclodiphosphate synthase (IspF). A two-step recombination strategy was used to construct ispF deletion mutants in M. tuberculosis but only wild-type double crossover strains were isolated. The chromosomal copy could be deleted when a second functional copy was provided on an integrating plasmid, demonstrating that ispF is an essential gene under the conditions tested thereby confirming its potential as a drug target. We attempted structure determination of the M. tuberculosis enzyme (MtIspF), but failed to obtain crystals. We instead analyzed the orthologue M. smegmatis IspF (MsIspF), sharing 73% amino acid sequence identity, at 2.2 A resolution. The high level of sequence conservation is particularly pronounced in and around the active site. MsIspF is a trimer with a hydrophobic cavity at its center that contains density consistent with diphosphate-containing isoprenoids. The active site, created by two subunits, comprises a rigid CDP-Zn2+ binding pocket with a flexible loop to position the 2C-methyl-D-erythritol moiety of substrate. Sequence-structure comparisons indicate that the active site and interactions with ligands are highly conserved. CONCLUSION: Our study genetically validates MtIspF as a therapeutic target and provides a model system for structure-based ligand design.