Reversible Denaturation of Brazil Nut 2S Albumin (Ber e1) and Implication of Structural Destabilization on Digestion by Pepsin
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- 作者：Stef J. Koppelman ; Willem F. Nieuwenhuizen ; Marco Gaspari ; Leon M. J. Knippels ; André ; H. Penninks ; Edward F. Knol ; Susan L. Hefle ; and Harmen H. J. de Jongh
- 刊名：Journal of Agricultural and Food Chemistry
- 出版年：2005
- 出版时间：January 12, 2005
- 年：2005
- 卷：53
- 期：1
- 页码：123 - 131
- 全文大小：296K
- 年卷期：v.53,no.1(January 12, 2005)
- ISSN：1520-5118
文摘
The high resistance of Brazil nut 2S albumin, previously identified as an allergen, against proteolysisby pepsin was examined in this work. Although the denaturation temperature of this protein exceedsthe 110 
C at neutral pH, at low pH a fully reversible thermal denaturation was observed at ~82 C.The poor digestibility of the protein by pepsin illustrates the tight globular packing. Chemical processing(i.e., subsequent reduction and alkylation of the protein) was used to destabilize the globular fold.Far-UV circular dichroism and infrared spectroscopy showed that the reduced and alkylated formhad lost its 
-structures, whereas the 
-helix content was conserved. The free energy of stabilizationof the globular fold of the processed protein as assessed by a guanidine titration study was only30-40% of that of the native form. Size exclusion chromatography indicated that the heavy chainlost its globular character once separated from the native 2S albumin. The consequences of thesechanges in structural stability for degradation by pepsin were analyzed using gel electrophoresisand mass spectrometry. Whereas native 2S albumin was digested slowly in 1 h, the reduced andalkylated protein was digested completely within 30 s. These results are discussed in view of thepotential allergenicity of Brazil nut 2S albumin.Keywords: Protein stability; protein folding; denaturation; pepsin; allergenicity
C at neutral pH, at low pH a fully reversible thermal denaturation was observed at ~82 C.The poor digestibility of the protein by pepsin illustrates the tight globular packing. Chemical processing(i.e., subsequent reduction and alkylation of the protein) was used to destabilize the globular fold.Far-UV circular dichroism and infrared spectroscopy showed that the reduced and alkylated formhad lost its -structures, whereas the -helix content was conserved. The free energy of stabilizationof the globular fold of the processed protein as assessed by a guanidine titration study was only30-40% of that of the native form. Size exclusion chromatography indicated that the heavy chainlost its globular character once separated from the native 2S albumin. The consequences of thesechanges in structural stability for degradation by pepsin were analyzed using gel electrophoresisand mass spectrometry. Whereas native 2S albumin was digested slowly in 1 h, the reduced andalkylated protein was digested completely within 30 s. These results are discussed in view of thepotential allergenicity of Brazil nut 2S albumin.Keywords: Protein stability; protein folding; denaturation; pepsin; allergenicity