In Silico and In Vitro Assessment of Portuguese Oyster (Crassostrea angulata) Proteins as Precursor of Bioactive Peptides
Abstract
:1. Introduction
2. Results and Discussion
2.1. Identified Proteins From C. angulata
2.2. In Silico Prediction of Potential Bioactivities
2.3. In Vitro Hydrolysis of Oyster Proteins
2.4. Confirmation of Bioactivities Through In Vitro Tests
2.4.1. ACE Inhibitory Activity
2.4.2. DPP-IV Inhibitory Activity
3. Materials and Methods
3.1. Materials
3.2. Oyster Meat Preparation
3.3. Proteomics Techniques and In Silico Analysis
3.3.1. Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis (SDS-Page) Analysis
3.3.2. Gel Slice and In-Gel Digestion
3.3.3. NanoLC–NanoESI–MS/MS Analysis
3.3.4. Mascot Database Search
3.3.5. BLAST Analysis of Oyster Proteins
3.4. In Vitro Analyses
3.4.1. Protein Isolation
3.4.2. Enzymatic Hydrolysis
3.4.3. Degree of Hydrolysis
3.4.4. Fractionation
3.4.5. Peptide Content Determination
3.4.6. Angiotensin-I Converting Enzyme (ACE) Inhibitory Activity Assay
3.4.7. Dipeptidyl Peptidase IV (DPP-IV) Inhibitory Activity Assay
3.4.8. Calculation of Inhibition Efficiency Ratio
3.5. Statistical Analysis
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Protein Name | Accession Number (NCBI) | Protein Score | Sequence Coverage (%) | Amino Acid Length | Molecular Weights from NCBI Database (kDa) | Molecular Weights Estimated from SDS-PAGE (kDa) |
---|---|---|---|---|---|---|
Myosin heavy chain, striated muscle isoform X1 * | XP_011442515.1 | 4670 | 48% | 1901 | 222.66 | 130.00 (A) |
82.50 (B) | ||||||
Paramyosin isoform X2 * | XP_011429256.1 | 2736 | 65% | 851 | 102.21 | 82.50 (B) |
Myosin heavy chain, striated muscle | EKC37566.1 | 789 | 20% | 2001 | 229.67 | 55.14 (C) |
44.55 (D) | ||||||
40.75 (E) | ||||||
11.66 (F) | ||||||
9.15 (G) | ||||||
5.76 (H) | ||||||
Actin * | EKC30049.1 | 1272 | 58% | 351 | 41.80 | 44.55 (D) |
Tropomyosin isoform X1 * | XP_019925727.1 | 579 | 70% | 251 | 33.02 | 40.75 (E) |
Hypothetical protein CGI_10010027 | EKC40031.1 | 220 | 27% | 151 | 20.58 | 11.66 (F) |
Myosin regulatory light chain B, smooth adductor muscle isoform X2 * | XP_011417566.1 | 112 | 50% | 151 | 18.63 | 9.15 (G) |
11.66 (F) | ||||||
5.76 (H) |
Protein Name | Number of Potential Bioactive Peptides | ||
---|---|---|---|
ACE Inhibitor | DPP-IV Inhibitor | Other Activities * | |
myosin heavy chain, striated muscle isoform X1 | 721 | 1147 | 395 |
paramyosin isoform X2 | 294 | 517 | 147 |
actin | 196 | 246 | 69 |
tropomyosin isoform X1 | 97 | 165 | 56 |
myosin regulatory light chain B, smooth adductor muscle isoform X2 | 83 | 104 | 39 |
Total | 1391 | 2179 | 706 |
Hydrolysate | Hydrolysis Conditions | Maximum DH (%) | Yield * (%) | Peptide Content (mg/mL) | |||
---|---|---|---|---|---|---|---|
E/S Ratio | Time (h) | pH | Temp. (°C) | ||||
PEH | 1:100 | 4 | 2 | 37 | 22.20 ± 0.97 a | 84.69 | 2.42 ± 0.06 a |
BRH | 1:100 | 4 | 7 | 50 | 17.86 ± 0.08 b | 35.14 | 1.53 ± 0.03 b |
PAH | 1:100 | 4 | 7 | 65 | 18.57 ± 0.61 b | 27.79 | 2.28 ± 0.03 a |
Fractions | Peptide Content (mg/mL) | Yield * (%) | Inhibition Efficiency Ratio a (%/mg/mL) | |
---|---|---|---|---|
DPP-IV Inhibitory Peptides | ACE Inhibitory Peptides | |||
PEH | 2.42 ± 0.06 a | 84.69 | 16.86 | 24.92 |
F1 | 0.32 ± 0.06 c | 10.95 | 153.00 | 217.05 |
F2 | 0.45 ± 0.02 c | 2.05 | 123.26 | 147.60 |
F3 | 1.73 ± 0.00 b | 54.74 | 27.11 | 29.04 |
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Gomez, H.L.R.; Peralta, J.P.; Tejano, L.A.; Chang, Y.-W. In Silico and In Vitro Assessment of Portuguese Oyster (Crassostrea angulata) Proteins as Precursor of Bioactive Peptides. Int. J. Mol. Sci. 2019, 20, 5191. https://doi.org/10.3390/ijms20205191
Gomez HLR, Peralta JP, Tejano LA, Chang Y-W. In Silico and In Vitro Assessment of Portuguese Oyster (Crassostrea angulata) Proteins as Precursor of Bioactive Peptides. International Journal of Molecular Sciences. 2019; 20(20):5191. https://doi.org/10.3390/ijms20205191
Chicago/Turabian StyleGomez, Honey Lyn R., Jose P. Peralta, Lhumen A. Tejano, and Yu-Wei Chang. 2019. "In Silico and In Vitro Assessment of Portuguese Oyster (Crassostrea angulata) Proteins as Precursor of Bioactive Peptides" International Journal of Molecular Sciences 20, no. 20: 5191. https://doi.org/10.3390/ijms20205191