Acta Crystallographica Section F

Structural Biology and Crystallization Communications

Volume 63, Part 9 (September 2007)


crystallization communications



Acta Cryst. (2007). F63, 787-791    [ doi:10.1107/S1744309107039838 ]

Crystallization and identification of the glycosylated moieties of two isoforms of the main allergen Hev b 2 and preliminary X-ray analysis of two polymorphs of isoform II

D. Fuentes-Silva, G. Mendoza-Hernández, V. Stojanoff, L. A. Palomares, E. Zenteno, A. Torres-Larios and A. Rodríguez-Romero

Abstract: Latex from Hevea brasiliensis contains several allergenic proteins that are involved in type I allergy. One of them is Hev b 2, which is a [beta]-1,3-glucanase enzyme that exists in different isoforms with variable glycosylation content. Two glucanase isoforms were isolated from trees of the GV-42 clone by gel filtration, affinity and ion-exchange chromatography. Isoform I had a carbohydrate content of about 20%, with N-linked N-acetyl-glucosamine, N-acetyl-galactosamine, fucose and galactose residues as the main sugars, while isoform II showed 6% carbohydrate content constisting of N-acetyl-glucosamine, fucose, mannose and xylose. Both isoforms were crystallized by the hanging-drop vapour-diffusion method. Isoform I crystals were grown using 0.2 M trisodium citrate dihydrate, 0.1 M Na HEPES pH 7.5 and 20%(v/v) 2-propanol, but these crystals were not appropriate for data collection. Isoform II crystals were obtained under two conditions and X-ray diffraction data were collected from both. In the first condition (0.2 M trisodium citrate, 0.1 M sodium cacodylate pH 6.5, 30% 2-propanol), crystals belonging to the tetragonal space group P41 with unit-cell parameters a = b = 150.17, c = 77.41 Å were obtained. In the second condition [0.2 M ammonium acetate, 0.1 M trisodium citrate dihydrate pH 5.6, 30%(w/v) polyethylene glycol 4000] the isoform II crystals belonged to the monoclinic space group P21, with unit-cell parameters a = 85.08, b = 89.67, c = 101.80 Å, [beta] = 113.6°. Preliminary analysis suggests that there are four molecules of isoform II in both asymmetric units.

Keywords: Hevea brasiliensis; latex allergens; endo-[beta]-1,3-glucanases; plant glycans.

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