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The DOI matches its Crossref record; 26 of the 26 references it cites were found in a registry or an index.

Characterization of 2S albumin allergenic proteins for anaphylaxis in common buckwheat

Katsube-Tanaka, Tomoyuki, Monshi, Fakhrul Islam·2022·Food Chemistry: Molecular Sciences
DOI 10.1016/j.fochms.2022.100127PMC PMC9363963
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Abstract

2S albumin (2SA) is responsible for anaphylaxis following consumption of buckwheat in allergic individuals. To reduce allergen incidents, characterization of 2SA polypeptides is prerequisite, thus was analyzed in this study. Of the five 2S albumin genes (g03, g11, g13, g14, and g28), g03 was seemingly non-functional. The g14 content was 3- and 40-fold higher than that of g11/g28 and g13, respectively. The g11/g28 were more processed to a ∼8 kDa band from a 16 kDa band than g14 in seeds, agreeing with that g11/g28 have high similarity with Fag e 8kD. Meanwhile, anti-g13 produced only a single ∼10 kDa band. Modification of g13 and domain exchange between g13 and g14 suggested that the hydrophobicity of the first domain and the nature of some amino acids in g13 contributed, at least in part, to the lower apparent molecular weight of g13 than expected. Thus, g13 might be an unexplored and noteworthy allergen.

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Crossref · 10.1016/j.fochms.2022.100127

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Figures

Fig. 1 2S albumin genes identified in Buckwheat Genome DataBase (BGDB). 2S albumin genes (g000003.aua.1, g03; g000011.aua.1, g11; g000013.aua.1, g13; g000014.aua.1, g14; g000028.aua.1, g28) were indicated with red filled box. Arrows indicate the direction of the genes. Approximate distances between the genes were shown.
Fig. 1 2S albumin genes identified in Buckwheat Genome DataBase (BGDB). 2S albumin genes (g000003.aua.1, g03; g000011.aua.1, g11; g000013.aua.1, g13; g000014.aua.1, g14; g000028.aua.1, g28) were indicated with red filled box. Arrows indicate the direction of the genes. Approximate distances between the genes were shown.CC BY-NC-ND · PubMed Central
Fig. 2 Alignment for new alleles of the 2S albumin gene g03. Newly identified alleles of g03 (g03_s and g03_h) were compared with the registered alleles of g03 and g28 in the BGDB database. Translation initiation (ATG) of the g03 in the BGDB was corrected. Translation initiation, predicted termination (TGA, TAA), and deletion were highlighted with a black box. The position of 622 bp deletion was also indicated.
Fig. 2 Alignment for new alleles of the 2S albumin gene g03. Newly identified alleles of g03 (g03_s and g03_h) were compared with the registered alleles of g03 and g28 in the BGDB database. Translation initiation (ATG) of the g03 in the BGDB was corrected. Translation initiation, predicted termination (TGA, TAA), and deletion were highlighted with a black box. The position of 622 bp deletion was also indicated.CC BY-NC-ND · PubMed Central
Fig. 3 Immunoreactivity of the representative recombinant 2S albumin prepared in E. coli cells. (A) The representative alleles of 2S albumin genes (g11, g13, g14, g28) were expressed in E. coli cells. After disrupting the cells, soluble (S) and insoluble (P) proteins were separated and detected using SDS-PAGE with CBB staining. For a quantification, designated amounts of BSA were also applied. White and black triangles indicate the position of 2S albumin and BSA, respectively. (B) The reactivity of anti 2S albumin antibodies were examined against the recombinant 2S albumin prepared in E. coli cells (g11, g28, g13, g14).
Fig. 3 Immunoreactivity of the representative recombinant 2S albumin prepared in E. coli cells. (A) The representative alleles of 2S albumin genes (g11, g13, g14, g28) were expressed in E. coli cells. After disrupting the cells, soluble (S) and insoluble (P) proteins were separated and detected using SDS-PAGE with CBB staining. For a quantification, designated amounts of BSA were also applied. White and black triangles indicate the position of 2S albumin and BSA, respectively. (B) The reactivity of anti 2S albumin antibodies were examined against the recombinant 2S albumin prepared in E. coli cells (g11, g28, g13, g14).CC BY-NC-ND · PubMed Central
Fig. 4 Detection and quantification of 2S albumin in common buckwheat seeds. (A) Albumin fractions prepared from bulked seeds flour of cultivars ‘Shinano 1’ (S) and ‘Harunoibuki’ (H) were detected with CBB and antibodies against g11, g28, g13, and g14. Black and white triangles indicate the position of major and minor processed 2S albumin, respectively. Note that anti-g11 and anti-g28 were reactive with g11 and g28 and vice versa. (B) Accumulation levels of 2S albumin g11/g28, g13, and g14 were analyzed using appropriately diluted recombinant 2S albumin (STD 1–3, see Materials and Methods for the dilution ratio). Three randomly selected seeds from the wild type cultivar ‘Harunoibuki’ (WT) were examined. White and black triangles indicate the position of recombinant and native 2S albumin, respectively.
Fig. 4 Detection and quantification of 2S albumin in common buckwheat seeds. (A) Albumin fractions prepared from bulked seeds flour of cultivars ‘Shinano 1’ (S) and ‘Harunoibuki’ (H) were detected with CBB and antibodies against g11, g28, g13, and g14. Black and white triangles indicate the position of major and minor processed 2S albumin, respectively. Note that anti-g11 and anti-g28 were reactive with g11 and g28 and vice versa. (B) Accumulation levels of 2S albumin g11/g28, g13, and g14 were analyzed using appropriately diluted recombinant 2S albumin (STD 1–3, see Materials and Methods for the dilution ratio). Three randomly selected seeds from the wild type cultivar ‘Harunoibuki’ (WT) were examined. White and black triangles indicate the position of recombinant and native 2S albumin, respectively.CC BY-NC-ND · PubMed Central
Fig. 5 Comparison of apparent and calculated molecular weights of modified and domain-exchanged 2S albumin. (A) The four recombinant 2S albumin (g11r, g28r, g13r, g14r), N-terminus modified g13 (g13’), and g13/g14 domain exchange series from No.1 to No.8 were prepared in E. coli expression system and electrophoresed with buckwheat seed proteins (S) in SDS-PAGE. Black and white triangles indicate the position of 16 kDa and 8–10 kDa 2S albumin, respectively. (B) Molecular weight estimated from SDS-PAGE migration (n = 3) and that calculated from amino acid sequence were compared. The regression line was drawn for all the symbols except g13, g11, and g28 with a dotted line. Dashed line indicates y = x.
Fig. 5 Comparison of apparent and calculated molecular weights of modified and domain-exchanged 2S albumin. (A) The four recombinant 2S albumin (g11r, g28r, g13r, g14r), N-terminus modified g13 (g13’), and g13/g14 domain exchange series from No.1 to No.8 were prepared in E. coli expression system and electrophoresed with buckwheat seed proteins (S) in SDS-PAGE. Black and white triangles indicate the position of 16 kDa and 8–10 kDa 2S albumin, respectively. (B) Molecular weight estimated from SDS-PAGE migration (n = 3) and that calculated from amino acid sequence were compared. The regression line was drawn for all the symbols except g13, g11, and g28 with a dotted line. Dashed line indicates y = x.CC BY-NC-ND · PubMed Central

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Metadata

Authors
Katsube-Tanaka, T., & Monshi, F. I.
Venue
Food Chemistry: Molecular Sciences
Publisher
Elsevier BV
Published
2022-12
Volume
5
Pages
100127
ISSN
2666-5662
DOI
10.1016/j.fochms.2022.100127
PMC
PMC9363963
Cited by
4
References
26
Language
en
Licence
Funders
  • Japan Society for the Promotion of Science (20H02967)
  • Tojuro Iijima Foundation for Food Science and Technology

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