2V2Z

IspE in complex with ADP and CDPME

Summary for 2V2Z

• Entry DOI: 10.2210/pdb2v2z/pdb
• Related: 2V2Q 2V2V 2V2Y
• Descriptor: 4-DIPHOSPHOCYTIDYL-2C-METHYL-D-ERYTHRITOL KINASE, 4-DIPHOSPHOCYTIDYL-2-C-METHYL-D-ERYTHRITOL, ADENOSINE-5'-DIPHOSPHATE, ... (6 entities in total)
• Functional Keywords: transferase, 4-diphosphocytidyl-2c-methyl-d-erythritol, aquifex aeolicus, nucleotide-binding, isoprene biosynthesis, kinase, atp-binding, non-mevalonate
• Biological source: AQUIFEX AEOLICUS
• Total number of polymer chains: 2
• Total formula weight: 62492.32

Authors

Sgraja, T.,Alphey, M.S.,Hunter, W.N. (deposition date: 2007-06-08, release date: 2007-06-19, Last modification date: 2024-05-08)

Primary citation

Sgraja, T.,Alphey, M.S.,Ghilagaber, S.,Marquez, R.,Robertson, M.N.,Hemmings, J.L.,Lauw, S.,Rohdich, F.,Bacher, A.,Eisenreich, W.,Illarionova, V.,Hunter, W.N.
Characterization of Aquifex Aeolicus 4-Diphosphocytidyl-2C-Methyl-D-Erythritol Kinase -Ligand Recognition in a Template for Antimicrobial Drug Discovery.
FEBS J., 275:2779-, 2008

PubMed Abstract: 4-Diphosphocytidyl-2C-methyl-D-erythritol kinase (IspE) catalyses the ATP-dependent conversion of 4-diphosphocytidyl-2C-methyl-D-erythritol (CDPME) to 4-diphosphocytidyl-2C-methyl-d-erythritol 2-phosphate with the release of ADP. This reaction occurs in the non-mevalonate pathway of isoprenoid precursor biosynthesis and because it is essential in important microbial pathogens and absent from mammals it represents a potential target for anti-infective drugs. We set out to characterize the biochemical properties, determinants of molecular recognition and reactivity of IspE and report the cloning and purification of recombinant Aquifex aeolicus IspE (AaIspE), kinetic data, metal ion, temperature and pH dependence, crystallization and structure determination of the enzyme in complex with CDP, CDPME and ADP. In addition, 4-fluoro-3,5-dihydroxy-4-methylpent-1-enylphosphonic acid (compound 1) was designed to mimic a fragment of the substrate, a synthetic route to 1 was elucidated and the complex structure determined. Surprisingly, this ligand occupies the binding site for the ATP alpha-phosphate not the binding site for the methyl-D-erythritol moiety of CDPME. Gel filtration and analytical ultracentrifugation indicate that AaIspE is a monomer in solution. The enzyme displays the characteristic alpha/beta galacto-homoserine-mevalonate-phosphomevalonate kinase fold, with the catalytic centre positioned in a deep cleft between the ATP- and CDPME-binding domains. Comparisons indicate a high degree of sequence conservation on the IspE active site across bacterial species, similarities in structure, specificity of substrate recognition and mechanism. The biochemical characterization, attainment of well-ordered and reproducible crystals and the models resulting from the analyses provide reagents and templates to support the structure-based design of broad-spectrum antimicrobial agents.

PubMed: 18422643
DOI: 10.1111/J.1742-4658.2008.06418.X

Experimental method

X-RAY DIFFRACTION (2.25 Å)