Investigation of Se-containing proteins in Bertholletia excelsa H.B.K. (Brazil nuts) by ICPMS, MALDI-MS and LC–ESI-MS methods

Owing to recent studies which showed that Se supplementation in the diet can reduce the risk of several forms of cancer, efforts have been directed to identify anticarcinogenic activity in Se compounds from natural sources with the objective of using those as food supplements. As a part of a continuing research project directed at identifying Se-containing species in food stuffs, the distribution of selenium-containing proteins in water-soluble protein fraction of seeds of Bertholletia excelsa H.B.K., which typically have a high Se content, was studied under non-denaturing conditions and the effectiveness of protein fractional precipitation by ammonium sulfate was investigated with the objectives to preconcentrate the protein(s) of interest and to reduce the matrix complexity. By SEC–ICPMS, Se-containing proteins (selenoproteome with some additional proteins) were found demonstrating the usefulness of ICPMS as an “on-line assay” in sub-proteomics investigations. Fractions with Se-containing proteins, collected from SEC, were tryptically digested and tryptic digests were analyzed by MALDI-TOF-reflectron MS and capLC–ESI-QTOF-MS. Isotope patterns which are different from the typical isotope patterns of peptides containing C, H, O, N and S were used to identify selenium-containing peptides in the digests. Observed isotope patterns were slightly different from the predicted isotope patterns and partial deamidation of selenium-containing peptides is suggested to explain the modified isotope pattern. To our knowledge, this is the first report of the effect of partial deamidation of selenium-containing peptides on the observed selenium isotope pattern, although the evidence is indirect. According to the SEC–ICPMS study, Se-containing proteins in water-soluble protein fraction of Bertholletia excelsa H.B.K. seeds can be divided into two main sub-groups-high molecular weight Se-containing proteins and low molecular weight Se-containing proteins. LC–ESI-MS and MS/MS analysis of tryptic digest of low-molecular weight Se-containing proteins identified 2S albumins, which are rich in methionines; therefore, with a high probability of non-specific selenomethionine incorporation—as the proteins present in that fraction.

Isotope pattern changes showing the effect of deamidation from the normal form (a), C80H135N19O20Se, to the deamidated form (b), C80H134N18O21Se.Figure optionsDownload high-quality image (105 K)Download as PowerPoint slideResearch highlights▶ Importance of ICPMS as an “on-line assay” in selenoproteomics research. ▶ Distribution of Se-containing proteins in water-soluble protein fraction of Brazil nuts. ▶ Slightly modified Se-isotope pattern owing to the occurrences of partial deamidation. ▶ Identified proteins S14946, CAA00789, A25802, BAA96554, Q9LRC2, S14947, Q84ND2_BEREX.