Table 3.
Overview of previously verified and newly predicted emulsifier and antioxidant peptides from identified and abundant proteins in the three E. denticulatum extracts. Peptides are grouped by their predicted mode of function and listed with their annotated name, predicted score, parent protein(s) annotation, amino acid sequence, length (L), charge at pH 7 (z) as well as the abundance by length-normalized, relative intensity (ILrel) by semi-specific analysis for the three enzymatic extracts (Viscozyme® (V), Alcalase® (A), and Shearzyme®+Celluclast® (S + C), respectively).
| Type3 |
Annotation |
Score4 |
Protein(s)5 |
Sequence |
L |
z6 |
ILrel (V) |
ILrel (A) |
ILrel (S + C) |
|---|---|---|---|---|---|---|---|---|---|
| Verified emulsifier peptides1 | |||||||||
| α | 80-S-A* | 3.3 | c7052_g1_i1, c7052_g1_i2 | IGYTVRNSLRVTVRDLSNLGLILDALVR | 28 | 2 | 12.9% | 24.5% | 9.6% |
| 81-S-A | 2.9 | c7052_g1_i1 | AVKDAVRRATLLTKAAGTGLGKVLS | 25 | 4 | 8.6% | 14.6% | 6.9% | |
| β | 83-S-B* | 3.18 | c7052_g1_i1, c7052_g1_i2 | RAGSNSLSRISFGISNEADLRDQAR | 25 | 1 | 12.9% | 24.5% | 9.6% |
| 84-S-B* | 2.75 | c6313_g1_i1 | VGFACSGSAQTYLSFEGDNTGRGEEEVAI | 29 | −4.1 | 1.3% | 5.7% | 1.1% | |
| 85-S-B | 3.27 | c7052_g1_i1 | LSIREGGRSTGGFSAQVRAR | 20 | 3 | 8.6% | 14.6% | 6.9% | |
| 86-S-B* | 5.43 | c7052_g1_i1 | ELQVSARVTLEIEL | 14 | −2 | 8.6% | 14.6% | 6.9% | |
| γ | 87-S-G | 5.72 | c1505_g2_i1 | RELQRDDNVRNVRILLSSLVLLLDWLVCLL | 30 | −0.1 | 27.4% | 30.8% | 26.4% |
| 88-S-G | 5.06 | c1505_g2_i1 | AVLVVCLQQVRELQRDDNVRN | 21 | −0.1 | 27.4% | 30.8% | 26.4% | |
| Predicted emulsifier peptides | |||||||||
| α | pEa-1 | 3.77 | c1545_g1_i1 | VKRISGLIYEETRNVLKVF | 19 | 2 | 2.8% | 0.0% | 7.4% |
| pEa-2 | 3.64 | c13559_g1_i1 c6825_g1_i1 | IYKVLKQV | 8 | 2 | 2.2% | 0.0% | 8.1% | |
| pEa-3 | 3.62 | c10861_g1_i1 | RFFLRVVRGVRQKV | 14 | 5 | 0.5% | 3.7% | 0.6% | |
| pEa-4 | 3.44 | c14987_g1_i1** | VQKLSRVID | 9 | 1 | 0.0% | 0.0% | 0.0% | |
| pEa-5 | 3.37 | c6825_g1_i1 | VADLFERIASEAAKL | 15 | −1 | 2.2% | 0.0% | 8.1% | |
| β | pEb-1 | 4.80 | c7216_g1_i1 | VRIRVDCK | 8 | 1.9 | 2.5% | 5.4% | 2.8% |
| pEb-2 | 4.12 | c14987_g1_i1** | RLNCKID | 7 | 0.9 | 0.0% | 0.0% | 0.0% | |
| pEb-3 | 3.72 | c4419_g1_i1 | LKVELNSGGQMR | 12 | 1 | 1.7% | 0.0% | 2.0% | |
| pEb-4 | 3.55 | c907_g1_i1 | NIQKESTLHLVLRLRGGL | 18 | 2.1 | 15.1% | 5.3% | 6.6% | |
| pEb-5 | 3.20 | c13533_g1_i1 | AKVRVTC | 7 | 1.9 | 2.2% | 0.0% | 4.3% | |
| γ | pEg-1 | 5.20 | c13533_g1_i1 | ITTVLALVCVITQVMQASANEQEHVHEHEH | 30 | −3.7 | 2.2% | 0.0% | 4.3% |
| pEg-2 | 5.11 | c24_g1_i1.p2 | ILLVLCLSWLRRKVCRNHR | 19 | 5 | 0.0% | 3.4% | 0.2% | |
| pEg-3 | 5.10 | c10861_g1_i1 | EREYELQKEFATLVLAVV | 18 | −2 | 0.5% | 3.7% | 0.6% | |
| pEg-4 | 4.69 | c17933_g1_i1 | FLLVFFFFFTDEDT | 14 | −3 | 5.2% | 13.5% | 6.2% | |
| pEg-5 | 4.43 | c907_g1_i1 | QDQQRLIFA | 9 | 0 | 15.1% | 5.3% | 6.6% | |
| Verified antioxidant peptides2 | |||||||||
| SCA | 120-S-SCA | 0.49 | c6313_g1_i1 | RYVWN | 5 | 1 | 1.3% | 5.7% | 1.1% |
| 123-S-SCA* | 0.38 | c6313_g1_i1 | DFPVR | 5 | 0 | 1.3% | 5.7% | 1.1% | |
| 124-S-SCA* | 0.43 | c17304_g1_i1 | AGDWLIGDR | 9 | −1 | 9.8% | 3.9% | 2.3% | |
| Predicted antioxidant peptides | |||||||||
| SCA | pSCA-1 | 0.72 | c6963_g1_i1 | DFYAYIVFTWAGYHGVDLAKNKIASDF | 27 | −0.9 | 2.7% | 0.9% | 5.2% |
| pSCA-2 | 0.71 | c24_g1_i1 | WYHY | 4 | 0.1 | 0.0% | 3.4% | 0.2% | |
| pSCA-3 | 0.66 | c10861_g1_i1 | LQGSKFAVVEYGGIVDPILGLQP | 23 | −1 | 0.5% | 3.7% | 0.6% | |
| pSCA-4 | 0.64 | c13533_g1_i1 | ASANEQEHVHEHEHIIRTFS | 20 | −2.6 | 2.2% | 0.0% | 4.3% | |
| pSCA-5 | 0.64 | c4419_g1_i1 | VVPFSSWYAEQQRI | 14 | 0 | 1.7% | 0.0% | 2.0% | |
| CHE | pCHE-1 | 0.39 | c17933_g1_i1 | FFFFTDEDTFPSGPSLTTFCSP | 22 | −3.1 | 5.2% | 13.5% | 6.2% |
| pCHE-2 | 0.36 | c6405_g1_i2** | FAISLFRIFPASFMFMPFTH | 20 | 1.1 | 0.0% | 0.0% | 0.0% | |
| pCHE-3 | 0.35 | c6405_g1_i2** | PHPN | 4 | 0.1 | 0.0% | 0.0% | 0.0% | |
| pCHE-4 | 0.34 | c7216_g1_i1 | FFSALLLLMNFPSPTMSLCTDDD | 23 | −3.1 | 2.5% | 5.4% | 2.8% | |
| pCHE-5 | 0.34 | Multiple*** | HP | 2 | 0.1 | 4.2% | 0.1% | 9.9% | |
*Peptides were in their respective studies highlighted as particularly promising based on in vitro functional validation.
**c14987_g1_i1 and c6405_g1_i2 were only identified by a single peptide each and only through unspecific analysis (but in all samples) and the peptides are regarded as potential contaminants and thus protein identification as doubtful.
***The dipeptide can be found in five abundant proteins (c13559_g1_i1; c24_g1_i1; c6405_g1_i2; c6656_g1_i1; and c6825_g1_i1).
Verified in vitro emulsifying activity (Yesiltas et al., 2021).
Verified in vitro antioxidant activity (Submitted manuscript, Food Chemistry).
Type refers to mode of predicted function. For emulsifier peptides, α refers to peptides with predicted amphiphilic helical conformation, β refers to predicted amphiphilic sheet conformation, and γ refers to peptides with one end being mostly hydrophobic and the other mostly hydrophilic. For antioxidant peptides, SCA refers to predicted free radical scavengers while CHE refers to predicted metal chelators.
Predicted scores for emulsifier and antioxidant peptides were computed using EmulsiPred (García-Moreno et al., 2020, García-Moreno et al., 2020) and AnOxPePred (Olsen et al., 2020), respectively.
Protein annotation in accordance with the de novo protein database previously reported (Gregersen et al., 2020).
Net charge at pH = 7 was calculated using the peptide property calculator (Innovagen AB, Lund, Sweden).