Solution Structure and Stability against Digestion of rproBnIb, a Recombinant 2S Albumin from Rapeseed: Relationship to Its Allergenic Properties
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- 作者：David Pantoja-Uceda ; Oscar Palomares ; Marta Bruix ; Mayte Villalba ; Rosalí ; a Rodrí ; guez ; Manuel Rico ; and Jorge Santoro
- 刊名：Biochemistry
- 出版年：2004
- 出版时间：December 28, 2004
- 年：2004
- 卷：43
- 期：51
- 页码：16036 - 16045
- 全文大小：435K
- 年卷期：v.43,no.51(December 28, 2004)
- ISSN：1520-4995
文摘
NMR spectroscopy has been used to determine the solution structure of the precursor form ofthe recombinant napin BnIb, rproBnIb, a 2S albumin, 109-residue protein from the seeds of Brassicanapus. More than 90% of the side-chain proton resonances were unambiguously assigned from the analysisof two-dimensional correlation (COSY), total correlation (TOCSY), and nuclear Overhauser effect (NOESY)spectra. The final structures were computed by using restrained molecular dynamics on the basis of 1316upper-limit distance constraints derived from NOE cross-correlation intensities. The computed structuresexhibited a root-mean-square deviation (RMSD) radius of 0.66 Å for the backbone and 1.16 Å for theside-chain heavy atoms of the structural core. The resulting structure consists of five amphipathic helicesarranged in a right-handed super helix, a folding motif found in other proteins of the prolamin superfamily.As in the case of the mature protein, the recombinant precursor behaves as a plant food allergen. To traceout the origin and characteristics of its allergenic properties, rproBnIb was assayed against simulatedgastric fluid and found to be very resistant to proteolysis. Also, heat treatment of the protein followed upto 85 
C by circular dichroism showed a very limited unfolding, which was recovered after cooling to 20C, indicating a high thermal stability. These results suggest that rproBnIb, as other 2S albumins, may beable to reach the gut immune system intact. A comparison of the putative epitopes against IgE antibodiesof the three members of the prolamine family [2S albumins, nonspecific lipid transfer proteins (nsLTPs),and 
-amylase/trypsin inhibitors] indicates that there are not common surfaces of interaction with IgE.Though the epitopes appear to be located in different regions of the proteins, they do comply with therequirements of being solvent-exposed and flexible.
C by circular dichroism showed a very limited unfolding, which was recovered after cooling to 20C, indicating a high thermal stability. These results suggest that rproBnIb, as other 2S albumins, may beable to reach the gut immune system intact. A comparison of the putative epitopes against IgE antibodiesof the three members of the prolamine family [2S albumins, nonspecific lipid transfer proteins (nsLTPs),and -amylase/trypsin inhibitors] indicates that there are not common surfaces of interaction with IgE.Though the epitopes appear to be located in different regions of the proteins, they do comply with therequirements of being solvent-exposed and flexible.