title: Binding studies and structure determination of the recombinantly produced type-II 3-dehydroquinate dehydratase from Acinetobacter baumannii.
abstract: Dehydroquinase (3-dehydroquinate dehydratase, DHQD, EC 4.2.1.10) catalyzes the conversion of dehydroquinate to dehydroshikimate. DHQD from Acinetobacter baumannii (AbDHQD) was cloned, expressed and purified to homogeneity. The binding studies showed that two compounds quinic acid and citrazinic acid bound to AbDHQD at micromolar concentrations. AbDHQD was crystallized using 30% PEG-3350, 50mM tris-HCl and 1.0M MgSO4 at pH 8.0. Crystals of AbDHQD were stabilized with 25% glycerol for data collection at 100K. The X-ray intensity data were collected to 2.0Å resolution. Crystals belonged to monoclinic space group P21with cell dimensions, a=82.3, b=95.3, c=132.3Å and β=95.7°. The structure was solved with molecular replacement method and refined to values of 0.200 and 0.232 for Rcrystand Rfreefactors. The structures of 12 crystallographically independent molecules in the asymmetry unit were identical with r.m.s shifts for the Cαatoms ranging from 0.3Å to 0.8Å. They formed a dodecamer with four trimers arranged in a tetrahedral manner. The classical lid adopted an open conformation although a sulfate ion was observed in the substrate binding site. As a result of which, the compounds quinic acid and citrazinic acid could not bind to AbDHQD.
authors: Department of Biophysics, All India Institute of Medical Sciences, New Delhi, India.
authors: Naseer
authors: N
authors: Iqbal
authors: Department of Biophysics, All India Institute of Medical Sciences, New Delhi, India.
authors: Mukesh
authors: M
authors: Kumar
authors: Department of Biophysics, All India Institute of Medical Sciences, New Delhi, India.
authors: Pradeep
authors: P
authors: Sharma
authors: Department of Biophysics, All India Institute of Medical Sciences, New Delhi, India.
authors: Satya Prakash
authors: SP
authors: Yadav
authors: Department of Biophysics, All India Institute of Medical Sciences, New Delhi, India.
authors: Punit
authors: P
authors: Kaur
authors: Department of Biophysics, All India Institute of Medical Sciences, New Delhi, India.
authors: Sujata
authors: S
authors: Sharma
authors: Department of Biophysics, All India Institute of Medical Sciences, New Delhi, India. Electronic address: tpsingh.aiims@gmail.com.
authors: Tej P
authors: TP
authors: Singh
mesh_terms: D040981:Acinetobacter baumannii / Q000201:enzymology; D001426:Bacterial Proteins / Q000737:chemistry*; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D006836:Hydro-Lyases / Q000737:chemistry*; D008958:Models, Molecular; D011485:Protein Binding; D000072756:Protein Conformation, alpha-Helical; D020836:Protein Structure, Quaternary; D011801:Quinic Acid / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry
chemical_list: D001426:Bacterial Proteins; D011994:Recombinant Proteins; D011801:Quinic Acid; D006836:Hydro-Lyases; C020000:3-dehydroquinate dehydratase
keywords: Acinetobacter baumannii; Dehydroquinate dehydratase; Dodecamer; Flexible lid; Open conformation