Expanded bed adsorption as a fast technique for the large-scale purification of the complete isoform pool of Ber e 1, the major allergen from Brazil nuts
✍ Scribed by Evelien L. van Boxtel; Gerrit A. van Koningsveld; Stef J. Koppelman; Lambertus A. M. van den Broek; Alfons G. J. Voragen; Harry Gruppen
- Publisher
- John Wiley and Sons
- Year
- 2006
- Tongue
- English
- Weight
- 543 KB
- Volume
- 50
- Category
- Article
- ISSN
- 1613-4125
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✦ Synopsis
Abstract
A new, fast, large‐scale purification method for Ber e 1, the major allergen from Brazil nuts, using expanded bed adsorption (EBA) chromatography, is presented. Using EBA, crude extracts can be applied to a fluidized column, which allows the unhindered passage of particulate impurities, thereby avoiding time‐consuming centrifugation or filtration steps. With this new purification method, 2.8 g of Ber e 1 was obtained from 85 g defatted Brazil nut meal, essentially within 1 day. Various structural as well as immunochemical characteristics of the purified protein were determined, and compared to those of Ber e 1 purified using conventional chromatographic techniques. The complete pool of Ber e 1 isoforms was collected using EBA. The most abundant isoforms were observed to have pI around 8 and heterogeneity was observed in both the large and the small subunit of the heterodimeric protein. Ber e 1 has a highly ordered secondary structure. No apparent differences in immune reactivity were observed between EBA purified Ber e 1 and conventionally purified Ber e 1, using IgE‐binding experiments. Thus, using EBA, Ber e 1 can be purified fast and on gram‐scale, while having purity equal to that of conventionally purified Ber e 1.