In Silico Analysis of Edible Bird’s Nest Proteins as Potential Precursors for Bioactive Peptides (original) (raw)
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Peptides, 2014
Angiotensin-I-converting enzyme (ACE-I, EC 3.4.15.1), renin (EC 3.4.23.15), and dipeptidyl peptidase-IV (DPP-IV, EC 3.4.14.5) play key roles in the control of hypertension and the development of type-2 diabetes and other diseases associated with metabolic syndrome. The aim of this work was to utilize known in silico methodologies, peptide databases and software including Prot-Param (http://web.expasy.org/protparam/), Basic Local Alignment Tool (BLAST), ExPASy PeptideCutter (http://web.expasy.org/peptide cutter/) and BIOPEP (http://www.uwm.edu.pl/biochemia/index.php/pl/ biopep) to assess the release of potentially bioactive DPP-IV, renin and ACE-I inhibitory peptides from bovine and porcine meat proteins including hemoglobin, collagen and serum albumin. These proteins were chosen as they are found commonly in meat by-products such as bone, blood and low-value meat cuts. In addition, the bioactivities of identified peptides were confirmed using chemical synthesis and in vitro bioassays. The concentration of peptide required to inhibit the activity of ACE-I and DPP-IV by 50% was determined for selected, active peptides. Novel ACE-I and DPP-IV inhibitory peptides were identified in this study using both in silico analysis and a literature search to streamline enzyme selection for peptide production. These novel peptides included the ACE-I inhibitory tri-peptide Ile-Ile-Tyr and the DPP-IV inhibitory tri-peptide Pro-Pro-Leu corresponding to sequences f (182-184) and f (326-328) of both porcine and bovine serum albumin which can be released following hydrolysis with the enzymes papain and pepsin, respectively. This work demonstrates that meat proteins are a suitable resource for the generation of bioactive peptides and further demonstrates the usefulness of in silico methodologies to streamline identification and generation of bioactive peptides.
Journal of Proteomics, 2014
In the present study angiotensin I-converting enzyme (ACE) inhibitory peptides were isolated from egg-yolk protein preparation (YP). Enzymatic hydrolysis conducted using unconventional enzyme from Cucurbita ficifolia (dose: 1000 U/mg of hydrolyzed YP (E/S (w/w) = 1:7.52)) was employed to obtain protein hydrolysates. The 4-h hydrolysate exhibited a significant (IC 50 = 482.5 μg/mL) ACE inhibitory activity. Moreover, hydrolysate showed no cytotoxic activity on human and animal cell lines which makes it a very useful multifunctional method for peptide preparation. The compiled isolation procedure (ultrafiltration, size-exclusion chromatography and RP-HPLC) of bioactive peptides from YP hydrolysate resulted in obtaining peptides with the strong ACE inhibitory activity. One homogeneous and three heterogeneous peptide fractions were identified. The peptides were composed of 9-18 amino-acid residues, including mainly arginine and leucine at the N-terminal positions. To confirm the selected bioactive peptide sequences their analogs were chemically synthesized and tested. Peptide LAPSLPGKPKPD showed the strongest ACE inhibitory activity, with IC 50 value of 1.97 μmol/L.
Purification and identification of an ACE-inhibitory peptide from walnut protein hydrolysate
European Food Research and Technology, 2014
Three novel Angiotensin I-converting enzyme (ACE) inhibitory peptides were isolated from poultry viscera protein hydrolysate (PVPH). The purification steps involved ultrafiltration of PVPH to eliminate the high molecular weight peptides and RP-HPLC. The ACE inhibitory activity was resolved into three peaks on C 18 column with retention times of 15.0, 18.9 and 20.9 min and amino acid sequences of the three peptides were determined to be ARIYH, LRKGNLE and RVWCP, respectively. The peptides exhibited resistance to hydrolysis by gastrointestinal proteases and extreme temperature (100C) and pH (1-12) at least for 2 h. Two of these peptides were competitive inhibitors, while the third was noncompetitive. The in vitro ACE inhibitory activity of PVPH revealed it could be a promising economic source for nutraceutical applications. PRACTICAL APPLICATIONS Poultry viscera is a protein-rich by-product of poultry processing industry. The tissue protein and its hydrolysate offer huge potential for different bioactive peptides. In the present study three novel ACE inhibitory peptides were identified and characterized from poultry viscera protein hydrolysate (PVPH). These peptides could be tested further for their in vivo antihypertensive effect and as functional ingredient in foods.
In this study, soft-shelled turtle (Pelodiscus sinensis) egg white (SSTEW) proteins were digested by thermolysin and the resulting small peptides were further fractionated by reverse phase chromatography. Peptides with angiotensin I-converting enzyme inhibitory (ACEI) activity from these fractions were screened. A lysozyme-derived peptide, IW-11, from the fraction with the most effective ACEI was identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and its purified form showed effective ACEI activity in vitro (IC 50 = 4.39 ± 0.31 μM). The Lineweaver-Burk plots indicated that the inhibition towards ACE caused by this peptide is a competitive inhibition. The molecular docking study further revealed that the ACEI activity of IW-11 is mainly attributed to the formation of hydrogen bonds between the N-terminal residue of IW-11 and the S1 pocket (Ala354 and Tyr523) and the S2′ region (His513 and His353) of ACE. Moreover, the digestion parameters were further optimized and the target peptide (82% purity) was readily obtained (15% yield) without any cumbersome purification procedure. Notably, lysozyme C is the most abundant protein in SSTEW, which implies that an efficient production of this ACEI peptide from SSTEW is promising.
Malaysian Applied Biology
Blood cockle (Anadara granosa) is the most abundant and available bivalves in Malaysia. Blood cockles meat has high protein content and has potential to generate bioactive peptides. To date, no study has been reported on purification and identification of angiotensin I converting enzyme (ACE) inhibitory peptides from blood cockle meat. Thus, the objectives of this study were to purify and characterize ACE inhibitory peptide from blood cockle meat hydrolysate. ACE inhibitory peptides from blood cockle meat hydrolysate (CMH) were prepared by enzymatic protein hydrolysis using Protamex®. Crude CMH was characterized for its stability against gastrointestinal proteases, at varying pH (2–11) and temperature (4–90°C). Next, crude CMH was purified by ultrafiltration, ion exchange chromatography and reverse-phase chromatography and its amino acid sequence was identified. It was found that crude CMH was highly stable at low pH and temperature, and was resistant to gastrointestinal proteases (...
Biologically active peptides derived from egg proteins
World's Poultry Science Journal, 2013
During food processing, proteins are subjected to different modifications, of which most are the result of proteolytic enzyme activity. Due to these mild and easily controlled reaction conditions, this method of modification has been commonly applied in producing high quality food products. Processing affects the conformation of the molecule and its biological activity. Limited hydrolysis leads to improvement of both the functional and nutritional properties of proteins, as well as their organoleptic characteristics. Proteolytic enzymes release peptides that frequently show different specific biological activities. Bioactive peptides govern the function of other compounds in the bloodstream, digestive, immune and nervous systems. Food sources of bioactive peptides exhibiting anti-hypertensive, anti-tumour, antioxidant or antimicrobial activities include milk, eggs and fish. Biopeptides with specific biological activities can be used in pharmaceutical, cosmetic or food industries as ...
مجله دانشکده پزشکی اصفهان, 2014
Background: Due to the side effects of anti-hypertension drugs, extracting bioactive peptides from natural sources is of great importance. The mail goal of this study was identifying a peptide for enzyme hydrolytic of ostrich egg white protein hydrolysate (OEWPH) and investigating its inhibitory effects on angiotansin-1 converting enzyme (ACE). Methods: The ostrich egg white protein hydrolysate was prepared using pepsin and pancreatin and then fractionated using evaluated reveres-phase high-performance liquid chromatography (RP-HPLC). Tandem mass analysis of the purified peptide was used to reveal peptide sequence. ACE inhibitory effect and kinetic parameters of the reaction in the presence of the peptide was evaluated. Molecular docking was used to determine the interaction parameters of ACE-peptide comp. Findings: Peptide sequencing of the selected peptide revealed a DAESLSRLLG (MW = 1060/18 Da) and named DG-10. The DG-10 peptide showed a potent inhibitory effect on ACE with the h...
Winged bean [Psophocarpus tetragonolobus (L.) DC.] seed is a potential underexploited source of vegetable protein due to its high protein content. In the present work, undefatted and defatted winged bean seed hydrolysates, designated as UWBSH and DWBSH, respectively were produced separately by four proteolytic enzymes namely Flavourzyme, Alcalase, Bromelain, and Papain using pH-stat method in a batch reactor. Enzymatic hydrolysis was carried out over a period of 0.5 to 5 h. UWBSH and DWBSH produced were tested for their ACE inhibitory activity in relation to the hydrolysis time and degree of hydrolysis (DH). Maximum ACE inhibitory activity, both for UWBSH and DWBSH, were observed during 3 to 5 h of hydrolysis. Both, UWBSH (DH 91.84 %), and DWSBH (DH 18.72 %), produced by Papain at 5 h hydrolysis, exhibited exceptionally high ACE inhibitory activity with IC 50 value 0.064 and 0.249 mgmL −1 , respectively. Besides, papain-produced UWBSH and DWBSH were further frac-tionated into three fractions based on molecular weight (UWBSH-I, <10 kDa; UWBSH-II, <5 kDa; UWBSH-III, <2 kDa) and (DWBSH-I, <10 kDa; DWBSH-II, <5 kDa; DWBSH-III, <2 kDa). UWBSH-III revealed the highest ACE inhibitory activity (IC 50 0.003 mgmL −1) compared with DWBSH-III (IC 50 0.130 mgmL −1). The results of the present investigation revealed that winged bean seed hydro-lysates can be explored as a potential source of ACE inhibitory peptides suggesting their uses for physiological benefits as well as for other functional food applications. Keywords Undefatted and defatted winged bean seed. Proximate analysis. ACE inhibitory activity. IC 50 value. Degree of hydrolysis. pH-stat titration
Process Biochemistry, 2018
In this research, ovalbumin (OOW), one of the major components in ostrich egg white, was purified by anion exchange chromatography. Then, the purified OOW was subjected to alkaline hydrolysis (0.25 M NaOH) at 40°C for 2-10 h. The best angiotensin I-converting enzyme (ACE) inhibitory activity was observed at 8 h of hydrolysis. The OOW hydrolysate obtained at 8 h (8 h-hOOW) was purified by the reversed-phase high-performance liquid chromatography (RP-HPLC). The resulting peptide, YV, exhibited an IC 50 value of 63.97 μg/mL. Using a Lineweaver-Burk plot, YV was determined to be a competitive inhibitor, and the inhibition constant (K i) was found to be 55.20 μg/mL. The molecular docking analysis revealed that the binding between YV and the S1 and S2 pocket sites of ACE was mainly stabilized by a hydrogen bond. Moreover, YV maintained ACE inhibitory activity after gastrointestinal digestion and showed no cytotoxic effects on human red blood cells, human keratinocyte cells (HaCaT) and human lung fibroblasts cells (MRC-5). An in vitro test of intestinal absorption showed that YV had a high potential for absorption into the Caco-2 cell monolayer model. Therefore, these results suggest that the YV peptide can be applied for the development of novel natural antihypertensive products.
Characterization of edible bird’s nest by peptide fingerprinting with principal component analysis
Food Quality and Safety, 2017
OBJECTIVES: Proteins are the major component and play a key role in nutritious and therapeutic functions of edible bird's nest (EBN); however, limited studies have been conducted on the protein due to difficulties in extraction, isolation as well as identification. This study aimed to provide comprehensive information for the quality evaluation of EBN peptides, which would be a valuable reference for further study on EBN proteins. METHODS: Here, we developed a quality control method using high performance liquid chromatography (HPLC) peptide fingerprints deriving from EBN being digested with simulated gastric fluid. The characteristic peptide peaks were collected and identified by LC-MS/MS. RESULTS: The characteristic peptide peaks, corresponding to the protein fragments of acidic mammalian chitinase-like, lysyl oxidase, and Mucin-5AC-like, were identified and quantified. Interestingly, the principal component analysis indicated that the fingerprints were able to discriminate colour of EBN (white/red) and production sites (cave/house) of White EBN on the same weight basis. As proposed by the model developed in this study, Muc-5AC-like and AMCaselike proteins were the markers with the highest discriminative power. CONCLUSIONS: The overall findings suggest that HPLC peptide fingerprints were able to clearly demonstrate peptide profile differences between genuine and adulterated EBN samples; and classify EBN samples by its color and production site. In addition, the protein identification results suggested that Muc-5AC-like protein was the major protein in EBN.