Data in Brief 8 (2016) 123–131
Contents lists available at ScienceDirect
Data in Brief journal homepage: www.elsevier.com/locate/dib
Data Article
Data on the epitope mapping of soybean A2 and A3 glycinin Hanaa Saeed, Christine Gagnon, Elroy Cober, Steve Gleddie n Agriculture and Agri-Food Canada, Ottawa Research & Development Centre, Ottawa, Ontario K1A 0C6, Canada
a r t i c l e i n f o
abstract
Article history: Received 11 April 2016 Received in revised form 12 May 2016 Accepted 14 May 2016 Available online 21 May 2016
The data information provided in this article relate to our research article “Using patient serum to epitope map soybean glycinins reveals common epitopes shared with many legumes and tree nuts” (Saeed et al., 2016) [1]. Here we provide western blot detection of glycinin subunits by soy-sensitive human sera, ELISA screens with overlapping synthetic peptides (epitope mapping), and various database/server epitope searches. & 2016 Published by Elsevier Inc. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).
Keywords: Epitope mapping Soybean Glycinin Western blot
Specifications Table Subject area More specific subject area Type of data How data was acquired
n
Immunology Allergy Tables, Graphs, Figures Western blots were performed by screening total soy protein on 2D gels with soy-sensitive human sera and detecting with a secondary anti-IgE-HRP antibody. ELISAs were performed by screening a collection of synthetic peptides encompassing the glycinin sequences with soy-sensitive human sera. The IgE binding to the peptides was detected by a secondary anti-IgE-HRP antibody.
DOI of original article: http://dx.doi.org/10.1016/j.molimm.2015.12.008 Corresponding author. E-mail address: [email protected] (S. Gleddie).
http://dx.doi.org/10.1016/j.dib.2016.05.027 2352-3409/& 2016 Published by Elsevier Inc. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).
124
H. Saeed et al. / Data in Brief 8 (2016) 123–131
Data format Experimental factors Experimental features Data source location Data accessibility
Epitope sequence similarity searches were done using the SDAP website: (http://fermi.utmb.edu/) B-cell epitope predictions were done using the following servers: ABCpred (http://www.imtech.res.in/raghava/abcpred/) BepiPred 1.0 (http://www.cbs.dtu.dk/services/BepiPred/) SVMTriP (http://sysbio.unl.edu/SVMTriP/) Raw, analyzed Human serum samples were acquired from individuals that exhibited a sensitivity to soybean and to other legumes/nuts Western blot, ELISA (epitope mapping) Canada and USA Data is provided with this article
Value of the data
Better understanding of soy storage protein allergens may contribute to allergy management strategies.
It may also contribute to the generation of hypoallergenic soybean cultivars. Provide risk assessment tools for the evaluation and characterization of the allergenicity of novel foods.
1. Data The data presented here show the western blot detection of A2 or A3 subunits by soy-sensitive human sera (Fig. 1) and ELISA screens (Figs. 2 and 3) of these patient sera with overlapping synthetic peptides (Pepsets). Serum specificity is also confirmed by cross-screening the A2 Pepset with a serum that does not bind to the A2 cluster on western blot (Fig. 4). Also contained in this article is SDAP (Structural Database of Allergenic Proteins) sequence similarity search results (Tables 1 and 2) of the epitopes reported by Saeed et al. (2016) [1] and theoretical B-cell epitope prediction data on the full length sequences of A2 and A3 subunits (Table 3).
2. Experimental design, materials and methods 2.1. Patient serum Soy-sensitive human sera used in the western blots and epitope mapping are previously described [1]. 2.2. Immunoblot analysis Western blotting of human sera was conducted as previously described [2]. Membranes were hybridized with serum dilutions ranging from 1/50 to 1/500. 2.3. Epitope mapping Two peptide sets representing the mature amino acid sequences of glycinin A2 (P04405, 90 peptides) and A3 (BAB15802, 104 peptides) were synthesized and biotinylated by Mimotopes (http://
H. Saeed et al. / Data in Brief 8 (2016) 123–131
4
125
7
75 KDa 50 KDa 37 KDa 25 KDa 20 KDa 15 KDa 4
Patient 1
1/500
7
4
75 KDa
75 KDa
50 KDa
50 KDa
37 KDa
37 KDa
25 KDa 20 KDa
25 KDa 20 KDa
15 KDa
15 KDa
4
Patient 2 1/400
7
4
75 KDa
75 KDa
50 KDa
50 KDa
37 KDa
37 KDa
25 KDa 20 KDa
25 KDa 20 KDa
15 KDa
15 KDa
4
75 KDa 50 KDa
Patient 3 1/200
Patient 4 1/500
7
4
Patient 5 1/150
Patient 6 1/300
7
7
7
75 KDa 50 KDa
37 KDa
37 KDa
25 KDa 20 KDa
25 KDa 20 KDa
15 KDa
15 KDa
Fig. 1. 2D western blots of patient sera on soybean seed protein gels (bottom 6 panels). Top panel is a Coomassie-stained 2D gel illustrating the A1/A2 (red) and A3 (blue) glycinins.
126
H. Saeed et al. / Data in Brief 8 (2016) 123–131
Fig. 2. Epitope mapping of A2 and A3 Pepsets with patient serum 1–6. X-axis indicates Pepset peptide number and y-axis indicates ratio of colorimetric detection in the patient sample vs control sample.
www.mimotopes.com) via parallel array platform. Quality Control Assurance was provided for both peptide synthesis and biotinylation by reverse phase HPLC (RP-HPLC), and by mass spectrometry (MS) respectively. The biotinylated 12-mer peptides, frame-shifted by three residues were used as per manufacturer's instructions (Application/Method PT3013). DMSO was used to resuspend the dry peptides and streptavidin-coated high capacity plates (Pierce #15500) pre-blocked with SuperBlock™
H. Saeed et al. / Data in Brief 8 (2016) 123–131
127
Fig. 3. Epitope mapping of A2 and A3 Pepsets using pooled sera. X-axis indicates Pepset peptide number and y-axis indicates ratio of colorimetric detection in the patient sample vs. control sample.
Fig. 4. Serum specificity control. Screening of A2 Pepset with Patient 4 serum which only bound to A3 subunit on western blot (see Fig. 1). X-axis indicates Pepset peptide number and y-axis indicates ratio of colorimetric detection in the patient sample vs control sample.
buffer were used to capture the biotinylated peptides. Serum was diluted at 1/50 in TBS-BSA 2% except for Patients 4 (1/100) and 5 (1/50 or 1/100). The secondary mouse anti-human IgE-HRP (Southern Biotech, Birmingham, Alabama, #9160-05) was diluted at 1/4000 in TBS-BSA 2%. SureBlue Reserve TM TMB microwell peroxidase substrate (KPL, Gaithersburg, Maryland, #53-00-01) was added to the plate, the reaction was stopped by acidification and colorimetric detection was performed on a Tecan Sunrise microplate reader with Magellan™ data analysis software (Tecan group AG, Männedorf, Switzerland) at 450 nm. Each experiment was performed in duplicate. Negative controls were performed using the same protocol, but the addition of human sera was omitted. The data was normalized by calculating the ratio of experimental to negative control and graphed.
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Table 1 SDAP sequence similarity of A3 epitopes (food only).n Rank
Allergen
Source
PD index
Location
Matching region
1 2 3 4 5 6 9 10 12 14 15 16 18 20 21 22 29 1 2 3 4 6 7 8 11 12 14 16 18 19 21 22 23 24 25 1 2 3
A3.1 epitope A4 Vig r 2.0201 Tri a glutenin Pru du 6.01 Pru du 6.01 Pru du 6.02 A1a Pru du 6.01 Tri a gliadin Tri a gliadin Tri a gliadin Tri a gliadin Bra j 1 Bra r 1 Tri a gliadin β-conglycinin (α0 ) A3.2 epitope A4 Len c 1.0101 A1a A1b Len c 1.0102 Ara h 3 Vig r 2 β-conglycinin (α0 ) Pru du 6.01 A2 Vig r 2 β-conglycinin (α) Fag e 1 Jug r 4 Cor a 9 Car i 4 Ara h 1 A3.3 epitope A4 Lup an 1.0101
Soybean Soybean Mung bean Wheat Almond Almond Almond Soybean Almond Wheat Wheat Wheat Wheat Indian mustard Field mustard Wheat Soybean Soybean Soybean Lentil Soybean Soybean Lentil Peanut Mung bean Soybean Almond Soybean Mung bean Soybean Buckwheat English walnut Hazelnut Pecan Peanut Soybean Soybean Lupin
0.00 1.15 3.82 3.83 3.87 3.87 3.87 4.02 4.06 4.14 4.34 4.48 4.84 4.93 4.95 4.97 5.16 0.00 0.00 3.80 4.26 4.66 4.71 5.39 5.86 5.86 6.16 6.17 6.30 6.46 6.49 6.54 6.57 6.57 6.57 0.00 4.84 7.00
214–222 214–222 218–226 191–199 100–108 136–144 209–217 205–213 122–130 23–31 142–150 260–268 208–216 88–96 134–142 119–127 433–441 226–237 227–238 144–155 218–229 217–228 144–155 237–248 183–194 366–377 269–280 215–226 181–192 331–342 241–252 231–242 238–249 232–243 304–315 313–324 316–327 101–112
KQGQHQQQE KQGQHQQEE QQGQESQQE QQGQSQQQQ QQGRQQEQE (epitope HS#5) QQGRQQQEE (epitope HS#5) QQGRQQQQQ (epitope) QKGKHQQEE QQGQQEQQQ (epitope HS#5) QQQQQQQQE KQQQQQQQQ QQPQQQQQQ HQQQQQQQE QQGQQLQHE QQGQQQQMQ QQAQQQQQQ EQQQRQQQE GSVLSGFSKHFL GSVLSGFSKHFL PSFLSGFSKNIL GSILSGFTLEFL (epitope) GSILSGFAPEFL PSFLSGFNKSIL GNIFSGFTPEFL (epitope) QSYLQGFSKNIL QSYLQGFSKNIL NNVFSGFNTQLL SNILSGFAPEFL (A2.5 epitope) QSYLRGFSKNIL QSYLQGFSRNIL ANILSGFQDEIL NNVFSGFDADFL (epitope) NNVFSGFDAEFL (epitope) NNVFSGFDAEFL (epitope) SSYLQGFSRNTL EEEDQPRPDHPP EDEDKPRPSRPS EQEQQPRPQRRQ
n
Only scores up to 8 are indicated.
2.4. B-cell epitope prediction servers Three popular B-cell epitope prediction servers were tested with the A2 and A3 sequences. ABCpred server predicts B cell epitopes using a recurrent neural network (machine based technique) using fixed length patterns [3]. Lengths of epitopes varying from 10–16 amino acids were tested. BepiPred 1.0 server uses a combination of a hidden Markov model and a propensity scale method [4]. SVMTriP uses support vector machine integrating tri-peptide similarity and propensity scores [5], where epitope lengths varying from 10–20 amino acids were tested. In all cases, a higher score reflects a higher probability that a sequence is an epitope.
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Table 2 SDAP sequence similarity of A2 epitopes (food only).n Rank
Allergen
Source
PD index
Location
Matching region
1 4 5 6 8 9 10 11 12 15 16 17 18 19 20 21 24 25 26 27 28 29 31 34 35 37 38 39 40 42 43 46 1 4 6 8 9 10 11 12 13 14 15 16 17 18 20 21 22 23 24 25 1 2 5 6 9 11 12 13
A2.1 epitope A1a A1b Tri a gliadin Pru du 6.02 Pis v 5 Ara h 3 Ana o 2 Lup an 1 Cor a 9 Jug r 4 A4 A3 Ses i 7 Tri a gliadin Lup an 1 Car i 4 Jug r 4 Ber e 2 Car i 4 Lup an 1 A1a β-conglycinin (α0 ) A2 Tri a gliadin Tri a glutenin Vig r 2 Tri a gliadin Ana o 1 Ara h 6 Tri a gliadin Jug r 2 A2.2 epitope Ara h 3 A1b A1a Ses i 6 Pis v 5 Ana o 2 Cor a 9 Pis v2 Car i 4 Jug r 4 Ber e 2 Pru du 6.02 Pru du 6.01 Ses i 7 Pis v2 Sin a 2 Fag e 1 A3 A4 A2.3 epitope A1a A1b Ara h 3 Pis v2 Ber e 2 Pru du 6.02 Cor a 9
soybean soybean soybean wheat almond pistachio peanut ashew Lupine Hazelnut English walnut Soybean Soybean Sesame Wheat Lupine Pecan English walnut Brazil nut Pecan Lupine Soybean Soybean Soybean Wheat Wheat mung bean wheat cashew peanut wheat English walnut Soybean Soybean Soybean Soybean Sesame Pistachio Cashew Hazelnut Pistachio Pecan English walnut Brazil nut Almond Almond Sesame Pistachio White mustard Buckwheat Soybean Soybean Soybean Soybean Soybean Peanut Pistachio Brazil nut Almond Hazelnut
0.00 1.36 1.36 4.72 4.99 4.99 5.01 5.22 5.27 5.59 5.70 5.83 5.83 5.83 5.86 5.90 6.14 6.14 6.18 6.28 6.32 6.45 6.46 6.51 6.56 6.58 6.60 6.60 6.61 6.63 6.65 6.78 0.00 1.31 2.71 2.77 3.21 3.40 4.02 4.08 4.25 4.31 4.31 4.58 4.70 4.70 4.73 5.84 6.61 6.94 7.07 7.07 0.00 0.00 1.22 2.21 4.67 4.92 4.92 5.10
121–129 124–132 121–129 241–249 137–145 126–134 138–146 118–126 126–134 138–146 210–218 129–137 129–137 141–149 134–142 51–59 131–139 130–138 128–136 210–218 583–591 115–123 147–155 109–117 210–218 191–199 330–338 92–100 42–50 54–62 137–145 111–119 130–141 124–135 130–141 133–144 144–155 135–146 127–138 147–158 151–162 140–151 139–150 137–148 146–157 193–204 150–161 146–157 181–192 161–172 138–149 138–149 136–153 139–156 136–153 133–150 157–174 143–160 152–169 153–170
QRPQDRHQK SRPQDRHQK SRPQDRHQK QQPQQQQQQ EDQQDRHQK SRFQDKHQK QQQQDSHQK GRFQDRHQK (2aa-epitope) QRPQSRREE RSEQDRHQK (epitope) RRQQQRQQR (epitope) QQLQDSHQK QQLQDSHQK RRFMDRHQK QQQQQQQQK QQPRPRQQE EFQQDRHQK (epitope) EFQQDRHQK (epitope) GRFQDQHQK HRRQQQHQQ (epitope) AQPQQQQQR QQPQQRGQS PRPHQPHQK QEPQESQQR QQQQQQEQK QQGQSQQQQ QREQQKQQE QQPQQQQQL QRQYDEQQK TRSSDQQQR QQQQQKQQQ QRGRDRQDP VHRFREGDLIAV VHRFDEGDLIAV IYHFREGDLIAV IYNFREGDLIAV VHRLRQGDIVAI IQRFRKGDIIAL IRRFRRGDIIAI IRHFREGDIIAL (5aa-epitope) VRHIREGDIIAL IRHFREGDIIAF IRHFREGDIIAF (5aa-epitope) VHHLKKGDIIAI IRHIREGDIIAL TRRIREGDVVAI VRQFRQGDILAL VRPIQEGDVIAL VEHVRHGDAIAM IFRIREGDVIPS IRHFNEGDVLVI IRHFNEGDVLVI GDLIAVPTGVAWWMYNNE GDLIAVPTGVAWWMYNNE GDLIAVPTGFAYWMYNNE GDLIAVPTGVAFWLYNDH GDIIALPAGVAHWIYNNG GDIIAIPAGVALWCYNDG GDIIALPAGVAYWSYNNG GDIIALPAGVAHWCYNDG
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Table 2 (continued ) Rank
Allergen
Source
PD index
Location
Matching region
14 15 16 18 19 20 21 22 23 1 3 4 7 8 10 11 12 13 14 15 16 17 19 20 22 25 26 1 3 7 8 9 10 11 13 14 22 33 39 41 45 46 1
Car i 4 Ana o 2 Pru du 6.01 Ses i 6 Jug r 4 Pis v 5 Ses i 7 A3 A4 A2.4 epitope A1b Ara h 3 Fag e 1 A1a Cor a 9 Car i 4 Pru du 6.02 Sin a 2 Jug r 4 A4 A3 Pru du 6.01 Ana o 2 Gly m Bd28K Pis v2 Ses i 7 Gal d vitellogenin (Gal d 6) A2.5 epitope Ara h 3 A1a A1b Pis v 5 Sin a 2 Ana o 1 A3 A4 Cap a 1 Gal d vitellogenin (Gal d 6) Gal d 6 Lyc e 4 Api g 2 Rub i 1 A2.6 epitope
Pecan Cashew Almond Sesame English walnut Pistachio Sesame Soybean Soybean Soybean Soybean Peanut Buckwheat Soybean Hazelnut Pecan Almond White mustard English walnut Soybean Soybean Almond Cashew Soybean Pistachio Sesame egg Soybean Peanut Soybean Soybean Pistachio White mustard Cashew Soybean Soybean Bell pepper egg Egg Tomato Celery Red raspberry Soybean
5.26 5.40 5.43 5.75 5.80 6.10 6.18 7.83 7.83 0.00 1.52 2.76 4.61 4.81 5.31 5.31 5.92 6.16 6.17 6.17 6.17 6.38 6.51 6.54 7.21 7.67 7.85 0.00 2.11 2.93 2.93 4.47 4.57 4.83 4.86 4.86 5.35 5.67 5.87 5.90 5.93 5.96 0.00
146–163 133–150 179–196 150–167 145–162 141–158 156–173 144–161 144–161 214–225 216–227 236–247 240–251 217–228 237–248 231–242 225–236 248–259 230–241 226–237 225–236 288–299 196–207 167–178 229–240 225–236 771–782 256–261 279–284 259–264 258–263 253–258 82–87 394–399 265–270 266–271 128–133 1395–1400 49–54 167–172 99–104 114–119 283–291
GDIIAFPAGVAHWCYNDG GDIIAIPAGVAHWCYNEG GDVVAIPAGVAYWSYNDG GDIVAIPSGAAHWCYNDG GDIIAFPAGVAHWSYNDG GDIIALPAGVANWCYNEG GDILALPAGLTLWFYNNG GDVLVIPPGVPYWTYNTG GDVLVIPPGVPYWTYNTG GSNILSGFAPEF GGSILSGFAPEF GGNIFSGFTPEF (epitope HS#2) GANILSGFQDEI GGSILSGFTLEF (epitope HS#2) GNNVFSGFDAEF (epitope HS#2) GNNVFSGFDAEF (5aa-epitope HS#2) GNNIFSGFDTQL (epitope HS#2) QQNILSGFDPQV GNNVFSGFDADF (epitope HS#2) GGSVLSGFSKHF GGSVLSGFSKHF (A3.2 epitope) GNNVFSGFNTQL (epitope HS#2) GRNLFSGFDTEL SHSVLSGFEPAI SNNILSAFDEEI TKNIFNGFDDEI ANQILNSIAGQW KGGLRV RGGLRI (epitope HS#3) KGGLSV (epitope HS#3) KGGLSV KGDLQV KGGLYL KGGMSV EGGLSV EGGLSV PGSLRV TGGLQL RTGIRI ESGLHV KCGIRI KGGAEI QCVETDKGC
n
Only scores up to 8 are indicated.
Table 3 B-cell epitope prediction. Name
Amino acid
Sequence
ABCpred
Bepipred
SVMTriP
A3.1 A3.2 A3.3 A2.1 A2.2 A2.3 A2.4 A2.5 A2.6
214–222 226–237 313–324 121–129 130–141 136–153 214–225 256–261 283–291
KQGQHQQQE GSVLSGFSKHFL EEEDQPRPDHPP QRPQDRHQK VHRFREGDLIAV GDLIAVPTGVAWWMYNNE GSNILSGFAPEF KGGLRV QCVETDKGC
0.72 – 0.81 – 0.90 0.87 0.82 0.80 –
1.63 – 2.72 1.33 – – – – 1.03
– 0.737 – 0.293 0.428 0.319 0.384 0.302 0.299
(Rank 8) (Rank 7) (Rank (Rank (Rank (Rank
4) 7) 7) 9)
Amino acid number corresponds to position of full length sequence. A dash (–) indicates that no epitope was found. Scores are listed for the 3 different methods tested. Only the highest score is listed if the epitope was found in multiple lengths tested (10– 20 mer).
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Acknowledgment The authors would like to acknowledge the late Dr. Susan Hefle and Dr. Steve Taylor (University of Nebraska) for their generous gift of patient sera. Supported by Agriculture and Agri-Food Canada, and the Genomics Research and Development Initiative of the Government of Canada.
Appendix A. Transparency document Transparency document associated with this article can be found in the online version at http://dx. doi.org/10.1016/j.dib.2016.05.027.
References [1] H. Saeed, C. Gagnon, E.R. Cober, S. Gleddie, Using patient serum to epitope map soybean glycinins reveals common epitopes shared with many legumes and tree nuts, Mol. Immunol. 70 (2016) 125–133. [2] C. Gagnon, V. Poysa, E.R. Cober, S. Gleddie, Soybean allergens affecting North American patients identified by 2D gels and mass spectrometry, Food Anal. Methods 3 (2010) 363–374. http://dx.doi.org/10.1007/s12161-009-9090-3. [3] S. Saha, G.P.S. Raghava, Prediction of continous B-cell epitopes in an antigen using recurrent neural network, Proteins 65 (2006) 40–48. [4] J.E.P. Larsen, O. Lund, M. Nielson, Improved method for predicting linear B-cell epitopes, Immunome Res. 2 (2006) 2. [5] B. Yao, L. Zhang, S. Liang, C. Zhang, SVMTriP: a method to predict antigenic epitopes using support vector machine to integrate tri-peptide similarity and propensity, PLoS ONE 7 (2012) e45152.
Contents lists available at ScienceDirect
Data in Brief journal homepage: www.elsevier.com/locate/dib
Data Article
Data on the epitope mapping of soybean A2 and A3 glycinin Hanaa Saeed, Christine Gagnon, Elroy Cober, Steve Gleddie n Agriculture and Agri-Food Canada, Ottawa Research & Development Centre, Ottawa, Ontario K1A 0C6, Canada
a r t i c l e i n f o
abstract
Article history: Received 11 April 2016 Received in revised form 12 May 2016 Accepted 14 May 2016 Available online 21 May 2016
The data information provided in this article relate to our research article “Using patient serum to epitope map soybean glycinins reveals common epitopes shared with many legumes and tree nuts” (Saeed et al., 2016) [1]. Here we provide western blot detection of glycinin subunits by soy-sensitive human sera, ELISA screens with overlapping synthetic peptides (epitope mapping), and various database/server epitope searches. & 2016 Published by Elsevier Inc. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).
Keywords: Epitope mapping Soybean Glycinin Western blot
Specifications Table Subject area More specific subject area Type of data How data was acquired
n
Immunology Allergy Tables, Graphs, Figures Western blots were performed by screening total soy protein on 2D gels with soy-sensitive human sera and detecting with a secondary anti-IgE-HRP antibody. ELISAs were performed by screening a collection of synthetic peptides encompassing the glycinin sequences with soy-sensitive human sera. The IgE binding to the peptides was detected by a secondary anti-IgE-HRP antibody.
DOI of original article: http://dx.doi.org/10.1016/j.molimm.2015.12.008 Corresponding author. E-mail address: [email protected] (S. Gleddie).
http://dx.doi.org/10.1016/j.dib.2016.05.027 2352-3409/& 2016 Published by Elsevier Inc. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).
124
H. Saeed et al. / Data in Brief 8 (2016) 123–131
Data format Experimental factors Experimental features Data source location Data accessibility
Epitope sequence similarity searches were done using the SDAP website: (http://fermi.utmb.edu/) B-cell epitope predictions were done using the following servers: ABCpred (http://www.imtech.res.in/raghava/abcpred/) BepiPred 1.0 (http://www.cbs.dtu.dk/services/BepiPred/) SVMTriP (http://sysbio.unl.edu/SVMTriP/) Raw, analyzed Human serum samples were acquired from individuals that exhibited a sensitivity to soybean and to other legumes/nuts Western blot, ELISA (epitope mapping) Canada and USA Data is provided with this article
Value of the data
Better understanding of soy storage protein allergens may contribute to allergy management strategies.
It may also contribute to the generation of hypoallergenic soybean cultivars. Provide risk assessment tools for the evaluation and characterization of the allergenicity of novel foods.
1. Data The data presented here show the western blot detection of A2 or A3 subunits by soy-sensitive human sera (Fig. 1) and ELISA screens (Figs. 2 and 3) of these patient sera with overlapping synthetic peptides (Pepsets). Serum specificity is also confirmed by cross-screening the A2 Pepset with a serum that does not bind to the A2 cluster on western blot (Fig. 4). Also contained in this article is SDAP (Structural Database of Allergenic Proteins) sequence similarity search results (Tables 1 and 2) of the epitopes reported by Saeed et al. (2016) [1] and theoretical B-cell epitope prediction data on the full length sequences of A2 and A3 subunits (Table 3).
2. Experimental design, materials and methods 2.1. Patient serum Soy-sensitive human sera used in the western blots and epitope mapping are previously described [1]. 2.2. Immunoblot analysis Western blotting of human sera was conducted as previously described [2]. Membranes were hybridized with serum dilutions ranging from 1/50 to 1/500. 2.3. Epitope mapping Two peptide sets representing the mature amino acid sequences of glycinin A2 (P04405, 90 peptides) and A3 (BAB15802, 104 peptides) were synthesized and biotinylated by Mimotopes (http://
H. Saeed et al. / Data in Brief 8 (2016) 123–131
4
125
7
75 KDa 50 KDa 37 KDa 25 KDa 20 KDa 15 KDa 4
Patient 1
1/500
7
4
75 KDa
75 KDa
50 KDa
50 KDa
37 KDa
37 KDa
25 KDa 20 KDa
25 KDa 20 KDa
15 KDa
15 KDa
4
Patient 2 1/400
7
4
75 KDa
75 KDa
50 KDa
50 KDa
37 KDa
37 KDa
25 KDa 20 KDa
25 KDa 20 KDa
15 KDa
15 KDa
4
75 KDa 50 KDa
Patient 3 1/200
Patient 4 1/500
7
4
Patient 5 1/150
Patient 6 1/300
7
7
7
75 KDa 50 KDa
37 KDa
37 KDa
25 KDa 20 KDa
25 KDa 20 KDa
15 KDa
15 KDa
Fig. 1. 2D western blots of patient sera on soybean seed protein gels (bottom 6 panels). Top panel is a Coomassie-stained 2D gel illustrating the A1/A2 (red) and A3 (blue) glycinins.
126
H. Saeed et al. / Data in Brief 8 (2016) 123–131
Fig. 2. Epitope mapping of A2 and A3 Pepsets with patient serum 1–6. X-axis indicates Pepset peptide number and y-axis indicates ratio of colorimetric detection in the patient sample vs control sample.
www.mimotopes.com) via parallel array platform. Quality Control Assurance was provided for both peptide synthesis and biotinylation by reverse phase HPLC (RP-HPLC), and by mass spectrometry (MS) respectively. The biotinylated 12-mer peptides, frame-shifted by three residues were used as per manufacturer's instructions (Application/Method PT3013). DMSO was used to resuspend the dry peptides and streptavidin-coated high capacity plates (Pierce #15500) pre-blocked with SuperBlock™
H. Saeed et al. / Data in Brief 8 (2016) 123–131
127
Fig. 3. Epitope mapping of A2 and A3 Pepsets using pooled sera. X-axis indicates Pepset peptide number and y-axis indicates ratio of colorimetric detection in the patient sample vs. control sample.
Fig. 4. Serum specificity control. Screening of A2 Pepset with Patient 4 serum which only bound to A3 subunit on western blot (see Fig. 1). X-axis indicates Pepset peptide number and y-axis indicates ratio of colorimetric detection in the patient sample vs control sample.
buffer were used to capture the biotinylated peptides. Serum was diluted at 1/50 in TBS-BSA 2% except for Patients 4 (1/100) and 5 (1/50 or 1/100). The secondary mouse anti-human IgE-HRP (Southern Biotech, Birmingham, Alabama, #9160-05) was diluted at 1/4000 in TBS-BSA 2%. SureBlue Reserve TM TMB microwell peroxidase substrate (KPL, Gaithersburg, Maryland, #53-00-01) was added to the plate, the reaction was stopped by acidification and colorimetric detection was performed on a Tecan Sunrise microplate reader with Magellan™ data analysis software (Tecan group AG, Männedorf, Switzerland) at 450 nm. Each experiment was performed in duplicate. Negative controls were performed using the same protocol, but the addition of human sera was omitted. The data was normalized by calculating the ratio of experimental to negative control and graphed.
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Table 1 SDAP sequence similarity of A3 epitopes (food only).n Rank
Allergen
Source
PD index
Location
Matching region
1 2 3 4 5 6 9 10 12 14 15 16 18 20 21 22 29 1 2 3 4 6 7 8 11 12 14 16 18 19 21 22 23 24 25 1 2 3
A3.1 epitope A4 Vig r 2.0201 Tri a glutenin Pru du 6.01 Pru du 6.01 Pru du 6.02 A1a Pru du 6.01 Tri a gliadin Tri a gliadin Tri a gliadin Tri a gliadin Bra j 1 Bra r 1 Tri a gliadin β-conglycinin (α0 ) A3.2 epitope A4 Len c 1.0101 A1a A1b Len c 1.0102 Ara h 3 Vig r 2 β-conglycinin (α0 ) Pru du 6.01 A2 Vig r 2 β-conglycinin (α) Fag e 1 Jug r 4 Cor a 9 Car i 4 Ara h 1 A3.3 epitope A4 Lup an 1.0101
Soybean Soybean Mung bean Wheat Almond Almond Almond Soybean Almond Wheat Wheat Wheat Wheat Indian mustard Field mustard Wheat Soybean Soybean Soybean Lentil Soybean Soybean Lentil Peanut Mung bean Soybean Almond Soybean Mung bean Soybean Buckwheat English walnut Hazelnut Pecan Peanut Soybean Soybean Lupin
0.00 1.15 3.82 3.83 3.87 3.87 3.87 4.02 4.06 4.14 4.34 4.48 4.84 4.93 4.95 4.97 5.16 0.00 0.00 3.80 4.26 4.66 4.71 5.39 5.86 5.86 6.16 6.17 6.30 6.46 6.49 6.54 6.57 6.57 6.57 0.00 4.84 7.00
214–222 214–222 218–226 191–199 100–108 136–144 209–217 205–213 122–130 23–31 142–150 260–268 208–216 88–96 134–142 119–127 433–441 226–237 227–238 144–155 218–229 217–228 144–155 237–248 183–194 366–377 269–280 215–226 181–192 331–342 241–252 231–242 238–249 232–243 304–315 313–324 316–327 101–112
KQGQHQQQE KQGQHQQEE QQGQESQQE QQGQSQQQQ QQGRQQEQE (epitope HS#5) QQGRQQQEE (epitope HS#5) QQGRQQQQQ (epitope) QKGKHQQEE QQGQQEQQQ (epitope HS#5) QQQQQQQQE KQQQQQQQQ QQPQQQQQQ HQQQQQQQE QQGQQLQHE QQGQQQQMQ QQAQQQQQQ EQQQRQQQE GSVLSGFSKHFL GSVLSGFSKHFL PSFLSGFSKNIL GSILSGFTLEFL (epitope) GSILSGFAPEFL PSFLSGFNKSIL GNIFSGFTPEFL (epitope) QSYLQGFSKNIL QSYLQGFSKNIL NNVFSGFNTQLL SNILSGFAPEFL (A2.5 epitope) QSYLRGFSKNIL QSYLQGFSRNIL ANILSGFQDEIL NNVFSGFDADFL (epitope) NNVFSGFDAEFL (epitope) NNVFSGFDAEFL (epitope) SSYLQGFSRNTL EEEDQPRPDHPP EDEDKPRPSRPS EQEQQPRPQRRQ
n
Only scores up to 8 are indicated.
2.4. B-cell epitope prediction servers Three popular B-cell epitope prediction servers were tested with the A2 and A3 sequences. ABCpred server predicts B cell epitopes using a recurrent neural network (machine based technique) using fixed length patterns [3]. Lengths of epitopes varying from 10–16 amino acids were tested. BepiPred 1.0 server uses a combination of a hidden Markov model and a propensity scale method [4]. SVMTriP uses support vector machine integrating tri-peptide similarity and propensity scores [5], where epitope lengths varying from 10–20 amino acids were tested. In all cases, a higher score reflects a higher probability that a sequence is an epitope.
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Table 2 SDAP sequence similarity of A2 epitopes (food only).n Rank
Allergen
Source
PD index
Location
Matching region
1 4 5 6 8 9 10 11 12 15 16 17 18 19 20 21 24 25 26 27 28 29 31 34 35 37 38 39 40 42 43 46 1 4 6 8 9 10 11 12 13 14 15 16 17 18 20 21 22 23 24 25 1 2 5 6 9 11 12 13
A2.1 epitope A1a A1b Tri a gliadin Pru du 6.02 Pis v 5 Ara h 3 Ana o 2 Lup an 1 Cor a 9 Jug r 4 A4 A3 Ses i 7 Tri a gliadin Lup an 1 Car i 4 Jug r 4 Ber e 2 Car i 4 Lup an 1 A1a β-conglycinin (α0 ) A2 Tri a gliadin Tri a glutenin Vig r 2 Tri a gliadin Ana o 1 Ara h 6 Tri a gliadin Jug r 2 A2.2 epitope Ara h 3 A1b A1a Ses i 6 Pis v 5 Ana o 2 Cor a 9 Pis v2 Car i 4 Jug r 4 Ber e 2 Pru du 6.02 Pru du 6.01 Ses i 7 Pis v2 Sin a 2 Fag e 1 A3 A4 A2.3 epitope A1a A1b Ara h 3 Pis v2 Ber e 2 Pru du 6.02 Cor a 9
soybean soybean soybean wheat almond pistachio peanut ashew Lupine Hazelnut English walnut Soybean Soybean Sesame Wheat Lupine Pecan English walnut Brazil nut Pecan Lupine Soybean Soybean Soybean Wheat Wheat mung bean wheat cashew peanut wheat English walnut Soybean Soybean Soybean Soybean Sesame Pistachio Cashew Hazelnut Pistachio Pecan English walnut Brazil nut Almond Almond Sesame Pistachio White mustard Buckwheat Soybean Soybean Soybean Soybean Soybean Peanut Pistachio Brazil nut Almond Hazelnut
0.00 1.36 1.36 4.72 4.99 4.99 5.01 5.22 5.27 5.59 5.70 5.83 5.83 5.83 5.86 5.90 6.14 6.14 6.18 6.28 6.32 6.45 6.46 6.51 6.56 6.58 6.60 6.60 6.61 6.63 6.65 6.78 0.00 1.31 2.71 2.77 3.21 3.40 4.02 4.08 4.25 4.31 4.31 4.58 4.70 4.70 4.73 5.84 6.61 6.94 7.07 7.07 0.00 0.00 1.22 2.21 4.67 4.92 4.92 5.10
121–129 124–132 121–129 241–249 137–145 126–134 138–146 118–126 126–134 138–146 210–218 129–137 129–137 141–149 134–142 51–59 131–139 130–138 128–136 210–218 583–591 115–123 147–155 109–117 210–218 191–199 330–338 92–100 42–50 54–62 137–145 111–119 130–141 124–135 130–141 133–144 144–155 135–146 127–138 147–158 151–162 140–151 139–150 137–148 146–157 193–204 150–161 146–157 181–192 161–172 138–149 138–149 136–153 139–156 136–153 133–150 157–174 143–160 152–169 153–170
QRPQDRHQK SRPQDRHQK SRPQDRHQK QQPQQQQQQ EDQQDRHQK SRFQDKHQK QQQQDSHQK GRFQDRHQK (2aa-epitope) QRPQSRREE RSEQDRHQK (epitope) RRQQQRQQR (epitope) QQLQDSHQK QQLQDSHQK RRFMDRHQK QQQQQQQQK QQPRPRQQE EFQQDRHQK (epitope) EFQQDRHQK (epitope) GRFQDQHQK HRRQQQHQQ (epitope) AQPQQQQQR QQPQQRGQS PRPHQPHQK QEPQESQQR QQQQQQEQK QQGQSQQQQ QREQQKQQE QQPQQQQQL QRQYDEQQK TRSSDQQQR QQQQQKQQQ QRGRDRQDP VHRFREGDLIAV VHRFDEGDLIAV IYHFREGDLIAV IYNFREGDLIAV VHRLRQGDIVAI IQRFRKGDIIAL IRRFRRGDIIAI IRHFREGDIIAL (5aa-epitope) VRHIREGDIIAL IRHFREGDIIAF IRHFREGDIIAF (5aa-epitope) VHHLKKGDIIAI IRHIREGDIIAL TRRIREGDVVAI VRQFRQGDILAL VRPIQEGDVIAL VEHVRHGDAIAM IFRIREGDVIPS IRHFNEGDVLVI IRHFNEGDVLVI GDLIAVPTGVAWWMYNNE GDLIAVPTGVAWWMYNNE GDLIAVPTGFAYWMYNNE GDLIAVPTGVAFWLYNDH GDIIALPAGVAHWIYNNG GDIIAIPAGVALWCYNDG GDIIALPAGVAYWSYNNG GDIIALPAGVAHWCYNDG
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Table 2 (continued ) Rank
Allergen
Source
PD index
Location
Matching region
14 15 16 18 19 20 21 22 23 1 3 4 7 8 10 11 12 13 14 15 16 17 19 20 22 25 26 1 3 7 8 9 10 11 13 14 22 33 39 41 45 46 1
Car i 4 Ana o 2 Pru du 6.01 Ses i 6 Jug r 4 Pis v 5 Ses i 7 A3 A4 A2.4 epitope A1b Ara h 3 Fag e 1 A1a Cor a 9 Car i 4 Pru du 6.02 Sin a 2 Jug r 4 A4 A3 Pru du 6.01 Ana o 2 Gly m Bd28K Pis v2 Ses i 7 Gal d vitellogenin (Gal d 6) A2.5 epitope Ara h 3 A1a A1b Pis v 5 Sin a 2 Ana o 1 A3 A4 Cap a 1 Gal d vitellogenin (Gal d 6) Gal d 6 Lyc e 4 Api g 2 Rub i 1 A2.6 epitope
Pecan Cashew Almond Sesame English walnut Pistachio Sesame Soybean Soybean Soybean Soybean Peanut Buckwheat Soybean Hazelnut Pecan Almond White mustard English walnut Soybean Soybean Almond Cashew Soybean Pistachio Sesame egg Soybean Peanut Soybean Soybean Pistachio White mustard Cashew Soybean Soybean Bell pepper egg Egg Tomato Celery Red raspberry Soybean
5.26 5.40 5.43 5.75 5.80 6.10 6.18 7.83 7.83 0.00 1.52 2.76 4.61 4.81 5.31 5.31 5.92 6.16 6.17 6.17 6.17 6.38 6.51 6.54 7.21 7.67 7.85 0.00 2.11 2.93 2.93 4.47 4.57 4.83 4.86 4.86 5.35 5.67 5.87 5.90 5.93 5.96 0.00
146–163 133–150 179–196 150–167 145–162 141–158 156–173 144–161 144–161 214–225 216–227 236–247 240–251 217–228 237–248 231–242 225–236 248–259 230–241 226–237 225–236 288–299 196–207 167–178 229–240 225–236 771–782 256–261 279–284 259–264 258–263 253–258 82–87 394–399 265–270 266–271 128–133 1395–1400 49–54 167–172 99–104 114–119 283–291
GDIIAFPAGVAHWCYNDG GDIIAIPAGVAHWCYNEG GDVVAIPAGVAYWSYNDG GDIVAIPSGAAHWCYNDG GDIIAFPAGVAHWSYNDG GDIIALPAGVANWCYNEG GDILALPAGLTLWFYNNG GDVLVIPPGVPYWTYNTG GDVLVIPPGVPYWTYNTG GSNILSGFAPEF GGSILSGFAPEF GGNIFSGFTPEF (epitope HS#2) GANILSGFQDEI GGSILSGFTLEF (epitope HS#2) GNNVFSGFDAEF (epitope HS#2) GNNVFSGFDAEF (5aa-epitope HS#2) GNNIFSGFDTQL (epitope HS#2) QQNILSGFDPQV GNNVFSGFDADF (epitope HS#2) GGSVLSGFSKHF GGSVLSGFSKHF (A3.2 epitope) GNNVFSGFNTQL (epitope HS#2) GRNLFSGFDTEL SHSVLSGFEPAI SNNILSAFDEEI TKNIFNGFDDEI ANQILNSIAGQW KGGLRV RGGLRI (epitope HS#3) KGGLSV (epitope HS#3) KGGLSV KGDLQV KGGLYL KGGMSV EGGLSV EGGLSV PGSLRV TGGLQL RTGIRI ESGLHV KCGIRI KGGAEI QCVETDKGC
n
Only scores up to 8 are indicated.
Table 3 B-cell epitope prediction. Name
Amino acid
Sequence
ABCpred
Bepipred
SVMTriP
A3.1 A3.2 A3.3 A2.1 A2.2 A2.3 A2.4 A2.5 A2.6
214–222 226–237 313–324 121–129 130–141 136–153 214–225 256–261 283–291
KQGQHQQQE GSVLSGFSKHFL EEEDQPRPDHPP QRPQDRHQK VHRFREGDLIAV GDLIAVPTGVAWWMYNNE GSNILSGFAPEF KGGLRV QCVETDKGC
0.72 – 0.81 – 0.90 0.87 0.82 0.80 –
1.63 – 2.72 1.33 – – – – 1.03
– 0.737 – 0.293 0.428 0.319 0.384 0.302 0.299
(Rank 8) (Rank 7) (Rank (Rank (Rank (Rank
4) 7) 7) 9)
Amino acid number corresponds to position of full length sequence. A dash (–) indicates that no epitope was found. Scores are listed for the 3 different methods tested. Only the highest score is listed if the epitope was found in multiple lengths tested (10– 20 mer).
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Acknowledgment The authors would like to acknowledge the late Dr. Susan Hefle and Dr. Steve Taylor (University of Nebraska) for their generous gift of patient sera. Supported by Agriculture and Agri-Food Canada, and the Genomics Research and Development Initiative of the Government of Canada.
Appendix A. Transparency document Transparency document associated with this article can be found in the online version at http://dx. doi.org/10.1016/j.dib.2016.05.027.
References [1] H. Saeed, C. Gagnon, E.R. Cober, S. Gleddie, Using patient serum to epitope map soybean glycinins reveals common epitopes shared with many legumes and tree nuts, Mol. Immunol. 70 (2016) 125–133. [2] C. Gagnon, V. Poysa, E.R. Cober, S. Gleddie, Soybean allergens affecting North American patients identified by 2D gels and mass spectrometry, Food Anal. Methods 3 (2010) 363–374. http://dx.doi.org/10.1007/s12161-009-9090-3. [3] S. Saha, G.P.S. Raghava, Prediction of continous B-cell epitopes in an antigen using recurrent neural network, Proteins 65 (2006) 40–48. [4] J.E.P. Larsen, O. Lund, M. Nielson, Improved method for predicting linear B-cell epitopes, Immunome Res. 2 (2006) 2. [5] B. Yao, L. Zhang, S. Liang, C. Zhang, SVMTriP: a method to predict antigenic epitopes using support vector machine to integrate tri-peptide similarity and propensity, PLoS ONE 7 (2012) e45152.
