Bovine Milk Proteins Research Articles

Real-time laser induced chemical derivatizations of peptide N-Terminus for in-situ mass spectrometric sequencing at sub-picomole and nanosecond scale

Distinguishing b- and y-ions is essential to compute amino acid sequences from either N- or C-terminus in mass spectrometry. We described herein a solvent free and real time on-plate derivatization approach that can tag N-terminus of peptides at microliter level with p-chlorobenzaldehyde or 2-hydroxy-5-methylisophthalaldehyde for matrix assisted laser desorption ionization mass spectrometry (MALDI MS). Less than 1 μL of sample solutions can be directly mixed with equal volumes of p-chlorobenzaldehyde or 2-hydroxy-5-methylisophthalaldehyde and α-cyano-4-hydroxycinnamic acid (CHCA), a matrix compound to co-crystalize with analytes for efficient absorption of laser energy and peptide ionization. When the mixture spotted on the sample plate is irradiated with the 3rd harmonic (355 nm) of Nd3+:YAG laser pulses (3 ns width), N-terminal amine groups of peptides instantly react with carbonyl groups of chlorobenzaldehyde or 2-hydroxy-5-methylisophthalaldehyde. Resultant peptides carrying with on-plate formed azomethine group (-CN-) are simultaneously protonated and isolated as precursor ions for subsequent collision-activated dissociation. The mass shift with unique Cl isotopic signature unambiguously distinguishes b ions from y ions and other ions. This method does not need extensive sample preparation and is useful for those samples with limited quantities down to sub-picomole level in sub-microliter volumes. The efficiency was demonstrated with synthetic peptides and tryptic peptides of model proteins. It was found that 2-hydroxy-5-methylisophthalaldehyde provides improved yield for peptides containing lysine residues. Unknown proteins of human saliva and bovine milk as well as phosphopeptides have been identified.

Orally administered milk-derived tripeptide improved cognitive decline in mice fed a high-fat diet.

The functions of the brain, which is thought of as an organ highly independent from the periphery, are often affected by the peripheral environment. Indeed, epidemiologic studies demonstrated that diabetes was a risk factor for dementia. It was also reported that the intake of dairy products, such as milk, reduces the risk of developing dementia. We found that mice on a short-term high-fat diet (HFD) for 1 wk had reduced cognitive function. Thus, using this acute model, we investigated the effects of milk-derived peptide on cognitive decline induced by HFD. Tyr-Leu-Gly (YLG), a tripeptide derived from αS1-casein, a major bovine milk protein, is released by gastrointestinal proteases. We found that orally administered YLG improved cognitive decline induced by 1-wk HFD intake in the object recognition test. YLG also improved cognitive decline in the object location test. Thus, we found that YLG improved cognitive decline induced by HFD. Next, we examined the effects of YLG on the hippocampus, a brain area essential for cognitive function. HFD intake decreased the number of 5-bromo-2'-deoxyuridine (BrdU)-positive cells, and this decrease was improved by YLG administration. HFD intake decreased nerve growth factor (NGF) and glial cell line-derived neurotrophic factor, whereas YLG increased NGF and ciliary neurotrophic factor, suggesting that these neurotropic factors play a role in hippocampal neurogenesis after YLG administration. In conclusion, we demonstrated that 1-wk HFD reduced cognitive function. Furthermore, we found that YLG, a milk-derived tripeptide, improved cognitive decline in mice on HFD. The HFD reduced neural stem cell proliferation, and YLG improved this reduction. YLG is the first reported milk peptide to improve cognitive decline induced by HFD intake.-Nagai, A., Mizushige, T., Matsumura, S., Inoue, K., Ohinata, K. Orally administered milk-derived tripeptide improved cognitive decline in mice fed a high-fat diet.

Effect of pathogens and somatic cell count on bovine milk proteins profile

Some nonthermal technologies have gained special interest as alternative approaches to thermal treatments. High pressure homogenization (HPH) and ultrasound (US) are two of the most promising approaches. They rely upon two different modes of action, although they share some mechanisms or ways of actions (mechanic burden against cells, cavitation and micronization, primary targets being the cell wall and the membrane, temperature and pressure playing important roles for their antimicrobial potential, and their effect on cells can be either positive or negative). HPH is generally used in milk and dairy products to break lipid micelles, whereas US is used for mixing and/or to obtain active compounds of food. HPH and US have been tested on pathogens and spoilers with different effects; thus, the main goal of this article is to describe how US and HPH act on biological systems, with a focus on antimicrobial activity, mode of action, positive effects, and equipment. The article is composed of three main parts: (i) an overview of US and HPH, with a focus on some results covered by other reviews (mode of action toward microorganisms and effect on enzymes) and some new data (positive effect and modulation of metabolism); (ii) a tentative approach for a comparative resistance of microorganisms; and (iii) future perspectives.High pressure homogenization (HPH) and ultrasound (US) can be used for food preservation.HPH and US exert a mechanic burden against microorganisms.HPH and US can be used for liquid foods.HPH and US are promising strategies, although further efforts are required.

Short communication: Processed bovine colostrum milk protein concentrate increases epithelial barrier integrity of Caco-2 cell layers

Colostrum plays an important role in initiating the development of the intestinal barrier in newborn mammals. Given its bioactivity, there is much interest in the potential use of bovine colostrum to improve human gastrointestinal health throughout the life span. There is evidence that bovine colostrum is effective at improving small intestinal barrier integrity and some indication that it may alter colonic motility. However, for colostrum to be used as a product to improve intestinal health, it needs to be bioactive after processing. The aim of this study was to determine whether industrial processing of bovine colostrum affects its ability to improve small intestinal barrier integrity or alter distal colon motility. Three colostrum sample types were compared; raw whole colostrum powder (WCP), raw skim colostrum powder (SCP), and industrially produced colostrum milk protein concentrate (CMPC). To determine whether these colostrum powders had different effects on small intestinal barrier integrity, their effects on the transepithelial electrical resistance across an in vitro intestinal epithelial layer (Caco-2 cells) were measured, both with and without a challenge from the proinflammatory cytokine tumor necrosis factor-α. These results showed that CMPC enhanced transepithelial electrical resistance across unchallenged epithelial cell layers, whereas the raw colostrum samples, WCP and SCP, did not have an effect. The colostrum samples were also compared to determine how they affect contractility in the distal colon isolated from the rat. Skim colostrum powder was the only sample to act directly on colonic tissue to modulate motility, increasing the amplitude of contractions. The results show that bovine colostrum is able to improve small intestinal barrier integrity and alter colon motility, and they implicate different components. The barrier integrity enhancement was apparent only in the industrial CMPC, which may have been due to the increase in protein concentration or the release of small peptides as a result of processing. The ability to alter colon motility was present in SCP but absent in WCP, again implying that an increase in protein concentration is responsible for the effect. However, this effect was not apparent for the industrially processed CMPC, suggesting denaturation or degradation of the active component. The beneficial effect of colostrum on small intestinal barrier integrity was present after processing, confirming that it is feasible to industrially produce an active product for gut health.

Alteration in propagating colonic contractions by dairy proteins in isolated rat large intestine

Gastrointestinal conditions in which the transit of contents is altered may benefit from nutritional approaches to influencing health outcomes. Milk proteins modulate the transit of contents along different regions, suggesting that they have varying effects on neuromuscular function to alter gastrointestinal motility. We tested the hypothesis that bovine whey and casein milk protein hydrolysates could have direct modulatory effects on colonic motility patterns in isolated rat large intestine. Casein protein hydrolysate (CPH), whey protein concentrate (WPC), whey protein hydrolysate (WPH), and a milk protein hydrolysate (MPH; a hydrolyzed blend of 60% whey to 40% casein) were compared for their effects on spontaneous contractile waves. These contractions propagate along the length of the isolated intact large intestine (22 cm) between the proximal colon and rectum and were detected by measuring activity at 4 locations. Milk proteins were perfused through the tissue bath, and differences in contraction amplitude and frequency were quantified relative to pretreatment controls. Propagation frequency was decreased by CPH, increased by MPH, and unaffected by intact whey proteins. The reduced motility with CPH and increased motility with MPH indicate a direct action of these milk proteins on colon tissue and provide evidence for differential modulation by hydrolysate type. These findings mirror actions on lower gastrointestinal transit reported in vivo, with the exception of WPH, suggesting that other factors are required.

Effects of thermal treatment on the physicochemical properties and osteogenic activity of lactoferrin

Lactoferrin as a key basic protein in bovine milk has attracted extensive attentions due to its multifunctional activities. Thermal treatment has been widely used during processing of dairy products, which might damage the bioactivity of heat-sensitive proteins. However, there has been little information on effect of heat treatment on the bioactivity of lactoferrin. In this study, the effects of four different thermal treatments (65°C/30 min, 72°C/10 s, 85°C/10 min, and 95°C/10 min) on lactoferrin were examined by sodium dodecyl sulfate–polyacrylamide gel electrophoresis, circular dichroism spectrum, electron microscope, dynamic light scattering, and semiquantitative RT-PCR. Results revealed that these treatments altered its second structure and surface hydrophobicity. Moreover, the osteogenic activity of lactoferrin decreased by analyzing the effect of different samples on the mitogen-activated protein kinase mRNA expression. These findings provided theory data for the research and development of lactoferrin as a functional component in food industry. Practical applications As we know, thermal treatment has been widely used during processing of dairy products. Lactoferrin is a key basic protein component in bovine milk and has a variety of biological activities such as osteogenic activity. However, very few studies have focused on the effect of heat treatment on the osteogenic activity of lactoferrin, thereby limiting its applications. The objective of the present study was to investigate the effects of heating on the structure and osteogenic activity of lactoferrin. Results showed that the regular heat-processing parameters in dairy industry and structural change of lactoferrin led to the loss of its osteogenic activity. These results will provide some theoretical and practical data for the development of lactoferrin as ingredient in functional food industry.

Characterization of the in situ pellicle ultrastructure formed under the influence of bovine milk and milk protein isolates

ObjectivesThe present study aimed to investigate if bovine milk or milk protein isolates, respectively, alter the ultrastructure of thein situ pellicle and might therefore have an influence on oral health. MethodsIn situ pellicle samples were formed on bovine enamel slabs exposed in the oral cavity of three subjects for 6, 30, 60 or 120 min. After 3 min of pellicle formation, mouthrinses were performed for 3 min with (non-)homogenized UHT- or fresh milk (0.3% or 3.8% fat), 30% UHT-treated cream or different types of casein- or milk protein isolates containing preparations. The specimens were removed after the exposure times and transmission electron microscopy (TEM) was performed. Native pellicle samples served as controls. ResultsTopical ultrastructural pellicle modifications were detected after mouthrinses with all types of homogenized UHT- or fresh milk and after the application of a 3% native casein micelles containing experimental solution. Atypical globular protein structures, identified as casein micelles, were temporarily adsorbed onto the pellicle. They were closely associated with lipid droplets. Furthermore, the mouthrinses occasionally affected the morphology of salivary bacteria. However, no notable ultrastructural alterations remained after 120 min of pellicle formation. ConclusionFor the first time, bovine milk- and micellar casein-induced pellicle modifications were revealed by TEM. The adsorption of micellar casein is possibly due to its molecular interactions. Clinical significanceBovine milk or micellar caseins provide some potential for the development of preventive strategies against bacterial biofilm formation or erosive processes at the tooth surface.

Strong Uranium(VI) Binding onto Bovine Milk Proteins, Selected Protein Sequences, and Model Peptides.

Hexavalent uranium is ubiquitous in the environment. In view of the chemical and radiochemical toxicity of uranium(VI), a good knowledge of its possible interactions in the environment is crucial. The aim of this work was to identify typical binding and sorption characteristics of uranium(VI) with both the pure bovine milk protein β-casein and diverse related protein mixtures (caseins, whey proteins). For comparison, selected model peptides representing the amino acid sequence 13-16 of β-casein and dephosphorylated β-casein were also studied. Complexation studies using potentiometric titration and time-resolved laser-induced fluorescence spectroscopy revealed that the phosphoryl-containing proteins form uranium(VI) complexes of higher stability than the structure-analog phosphoryl-free proteins. That is in agreement with the sorption experiments showing a significantly higher affinity of caseins toward uranium(VI) in comparison to whey proteins. On the other hand, the total sorption capacity of caseins is lower than that of whey proteins. The discussed binding behavior of milk proteins to uranium(VI) might open up interesting perspectives for sustainable techniques of uranium(VI) removal from aqueous solutions. This was further demonstrated by batch experiments on the removal of uranium(VI) from mineral water samples.

Elucidating the interaction mechanism of eriocitrin with β-casein by multi-spectroscopic and molecular simulation methods

Eriocitrin is a flavanone glycoside which exists in lemon or lime citrus fruits. It exhibits antioxidant, anti-cancer and anti-allergy activities. β-casein is important protein in bovine milk, under appropriate conditions, it can form stable micelle like structures in aqueous solution, which is beneficial to the nutritional ingredient transportation in body. In this work, the interaction mechanisms of eriocitrin with β-casein under physiological conditions (pH = 7.4) were investigated by utilizing multi-spectroscopic techniques and molecular docking methods. The endogenous fluorescence of β-casein was quenched by the interaction with eriocitrin and the quenching mode was static quenching. The interaction of eriocitrin with β-casein was a spontaneous reaction principally driven by hydrophobic interaction. At 310 K, the binding constant was equal to 6.68 × 105 L mol−1 and the number of binding sites was approximately equal to 1. The changes of β-casein conformation were confirmed by FTIR, circular dichroism spectroscopy and synchronous fluorescence spectroscopy. Moreover, molecular docking studies illustrated the most possible binding position of eriocitrin on β-casein.

Role of Bioactive Peptides in Reducing the Severity of Hypertension with the Inhibition of ACE

Bovine milk protein, fermented with LC (Lactobacillus casei) was used to evaluate its ACE inhibitory activity in-vitro. After evaluating the health status of animals by running hemto-chemical tests, four young dairy animals (22, 26, 30 and 34 months old) were selected on the basis of serum protein level for milk collection. Twenty-two months old animal was considered as control. After co-culturing of skim milk with LC for 24 h, peptide samples were purified via ultrafiltration by using Molecular weight cut-off; 10000 Da (PM-10) membrane. Protein hydrolysis was confirmed by applying ninhydrin reaction and SDS–PAGE (sodium dodecyl sulfate–polyacrylamide gel electrophoresis). Milk of 34-months old animal (D-34) showed greater ACE inhibitory activity in-vitro via ACE inhibitory assay. Moreover, D-34 was further analyzed for its peptides profile by using Q exactive Hybrid Quadrapole-Orbitrap Mass Spectrometry as shown in Graphical abstract. Peptides with higher concentration of Proline, Lysine and aromatic amino acid was identified and higher concentration of hydrophobic aliphatic amino acids in PIN-94 and PIN-332 indicated the usefulness of D-34 as ACE-inhibitor. On the basis of those hydrophobic amino acids it was concluded that Serum protein and milk protein level are interconnected and milk of middle-aged Animals with higher serum protein level is useful to inhibit ACE because with the growth of animal, serum protein contents also increases and those protein contents indicate the health status of the animal. Fermented milk peptides of middle-aged animals can be used as functional food and nutraceutical to reduce hypertension.

Protein profile of dairy products: Simultaneous quantification of twenty bovine milk proteins

While proteomic techniques allow the identification and relative quantification of thousands of proteins in a single run, methods for absolute quantification remain laborious. In this study, a newly developed multiple reaction monitoring (MRM) method using liquid chromatography mass spectrometry (LC-MS) that enables the simultaneous quantification of twenty key milk proteins is presented. The selected proteins comprise all individual caseins, the major whey proteins and most well-known milk fat globule membrane (MFGM) proteins. For validation, the twenty milk proteins in raw milk, raw cream, raw milk Emmental cheese and whey, were quantified as well as in eighteen commercial heat-treated dairy products. The method presented is ideally suited for various applications, for example, the comparison of the protein patterns in raw milk of cows at different stages of lactation or of different breeds.

Symposium review: Characterization of the bovine milk protein profile using proteomic techniques

Identification and characterization of the comprehensive bovine milk proteome has historically been limited due to the dichotomy of protein abundances within milk. The high abundance of a select few proteins, including caseins, α-lactalbumin, β-lactoglobulin, and serum albumin, has hindered intensive identification and characterization of the vast array of low-abundance proteins in milk due to limitations in separation techniques and protein labeling capacity. In more recent years, the development and advancement of proteomics techniques have yielded valuable tools for characterization of the protein profile in bovine milk. More extensive fractionation and enrichment techniques, including the use of combinations of precipitation techniques, immunosorption, gel electrophoresis, chromatography, ultracentrifugation, and hexapeptide-based binding enrichment, have allowed for better isolation of lower abundance proteins for further downstream liquid chromatography-tandem mass spectrometry approaches. The different milk subfractions isolated during these processes can also be analyzed as individual entities to assess the protein profile unique to the different fractions-for instance, investigation of the skim milk-associated proteome versus the milk fat globule membrane-associated proteome. Updates to high-throughput methods, equipment, and software have also allowed for greater interpretation and visualization of the data. For instance, labeling techniques have enabled analysis of multiplexed samples and more accurate comparison of specific protein abundances and quantities across samples, and integration of gene ontology analysis has allowed for a more in-depth and visual representation of potential relationships between identified proteins. Inclusively, these developments in proteomic techniques have allowed for a rapid increase in the number of milk-associated proteins identified and a better grasp of the relationships and potential functionality of the proteins within the milk proteome.

High-throughput quantitation of bovine milk proteins and discrimination of commercial milk types by external cavity-quantum cascade laser spectroscopy and chemometrics.

Analysis of bovine milk proteins is crucial in many food and non-food industrial applications, nevertheless labour-intensive wet-chemical, low-throughput methods are still routinely used. In this work, external cavity-quantum cascade laser (EC-QCL) mid-infrared spectroscopy is employed as a rapid method for protein analysis of commercial bovine milk. Combined analysis of the amide I and II bands enabled quantitation of individual proteins (casein, β-lactoglobulin, α-lactalbumin) and total protein content. IR spectra of spiked and diluted milk samples were employed for calibration of the target analytes in the presence of a complex matrix by partial least squares (PLS) regression modelling. A sample set of different milk types (pasteurized; differently processed extended shelf life, ESL; ultra-high temperature, UHT) was analysed, and results agreed well with reference methods. Quantitation of temperature sensitive proteins enables detailed distinction between milk types experiencing different heat loads during processing, and discrimination between diverse bovine milk types is successfully demonstrated.

Release of functional peptides from mother's milk and fortifier proteins in the premature infant stomach.

Digestion of milk proteins in the premature infant stomach releases functional peptides; however, which peptides are present has not been reported. Premature infants are often fed a combination of human milk and bovine milk fortifiers, but the variety of functional peptides released from both human and bovine milk proteins remains uncharacterized. This study applied peptidomics to investigate the peptides released in gastric digestion of mother’s milk proteins and supplemental bovine milk proteins in premature infants. Peptides were assessed for homology against a database of known functional peptides—Milk Bioactive Peptide Database. The peptidomic data were analyzed to interpret which proteases most likely released them from the parent protein. We identified 5,264 unique peptides from bovine and human milk proteins within human milk, fortifier or infant gastric samples. Plasmin was predicted to be the most active protease in milk, while pepsin or cathepsin D were predicted to be most active in the stomach. Alignment of the peptide distribution showed a different digestion pattern between human and bovine proteins. The number of peptides with high homology to known functional peptides (antimicrobial, angiotensin-converting enzyme-inhibitory, antioxidant, immunomodulatory, etc.) increased from milk to the premature infant stomach and was greater from bovine milk proteins than human milk proteins. The differential release of bioactive peptides from human and bovine milk proteins may impact overall health outcomes in premature infants.

Genetic Variation of β-Casein Gene Using AS-PCR and ARMS-PCR Techniques in Bovine Populations

Proteins in bovine milk are a common source of bioactive peptides. The peptides are released by the digestion of caseins and whey proteins. Therefore, the main aim of present study was ‎to evaluation of the frequency of beta-casein ‎variants of A1 (as bad peptide) and A2 (as ‎beneficial peptide) and also, A and B alleles in ‎Holstein and native breeds using both AS-PCR ‎and ARMS techniques. Totally, 40 Holstein, 38 Sarabi and 13 Gaja breeds were tested. In AS-PCR, three genotypes as A1A1, A1A2, and A2A2 were detected and frequencies of A1 allele of β-casein were calculated as 52.5, 51.31 and 50% in Holstein, Sarabi and Gaja breeds, respectively. Frequencies of heterozygote genotypes A1A2 were observed as 90, 71 and 100% in Holstein, Sarabi and Gaja breeds, in that order. Frequencies of A2A2 genotypes of β-casein were calculated as 2.5, 13.157, and 0.00% in the three studied breeds respectively. In ARMS-PCR, two genotypes as AA, and AB were observed and BB genotype wasn’t found. Frequencies of AA, and AB genotypes were as 80 and 20%, 94.73 and 5.26% and 100 and 0.00% in Holstein, Sarabi and Gaja populations, respectively. Subsequently, allele B frequencies were calculated as 3.38, 1.92, and 0.00% in order to these three breeds. Comparison of both techniques was showed that samples having A1A1 genotype in AS-PCR technique were as AB genotype in ARMS-PCR significantly (p ≥ 0.05). All of the identified alleles in both techniques weren’t in Hardy–Weinberg equilibrium. Overall, results of our study were revealed that frequencies of bad peptides were more than good peptide in the three different populations.

Identification of antihypertensive peptides in nutraceuticals by capillary electrophoresis-mass spectrometry

We present capillary electrophoresis-mass spectrometry (CE-MS) in combination with advanced chemometric tools for the analysis of bioactive compounds in food, in particular for the identification of antihypertensive peptides in a nutraceutical derived from a bovine milk protein hydrolysate. Different extracts of the nutraceutical were analyzed by CE-MS, and the electropherograms were processed using a novel data analysis workflow that included regions of interest (ROIs) compression and multivariate curve resolution alternating least squares (MCR-ALS). MCR-ALS permitted the description of the nutraceutical extract as ten characteristic components with their electrophoretic profiles and mass spectra. Twenty-two compounds were tentatively identified as antihypertensive bovine casein fragments through a mass search in a database of bioactive peptides. The identity of 17 antihypertensive peptides was reliably confirmed by capillary electrophoresis-tandem mass spectrometry. The proposed analytical approach demonstrated the potential to obtain a characteristic and activity-related fingerprint for quality control and authentication of the antihypertensive nutraceutical.

Digestive Responses to Fortified Cow or Goat Dairy Drinks: A Randomised Controlled Trial.

Fortified milk drinks are predominantly manufactured from bovine (cow) sources. Alternative formulations include those prepared with hydrolysed bovine milk proteins or from alternate bovidae species, such as caprine (goat) milk. Currently, there is little data on protein digestive and metabolic responses following ingestion of fortified milk drinks. To examine the digestive and metabolic responses to commercially-available fortified milks, young adults (n = 15 males: 15 females), in a randomised sequence, ingested isonitrogenous quantities of whole cow-protein (WC), whole goat-protein (WG), or partially-hydrolysed whey cow-protein (HC), commercial fortified milks. Plasma amino acid (AA) and hormonal responses were measured at baseline and again at 5 h after ingestion. Paracetamol recovery, breath hydrogen, and subjective digestive responses were also measured. Postprandial plasma AA was similar between WC and WG, while AA appearance was suppressed with HC. Following HC, there was a negative incremental AUC in plasma branched-chain AAs. Further, HC had delayed gastric emptying, increased transit time, and led to exaggerated insulin and GLP-1 responses, in comparison to whole protein formulas. Overall, WC and WG had similar protein and digestive responses with no differences in digestive comfort. Contrastingly, HC led to delayed gastric emptying, attenuated AA appearance, and a heightened circulating insulin response.

In vitro immunogenicity of various native and thermally processed bovine milk proteins and their mixtures

In vitro immunogenicity of various native and thermally processed (72°C/15 s and 100°C/30 s) bovine milk protein fractions, their mixtures, whey, and skim milk, was studied by analyzing the immune response of T helper (Th) cells in human peripheral blood mononuclear cells. The secretion of Th type cytokines induced by the protein stimulants was quantified while determining the heat-induced protein denaturation. Purified whey proteins, caseins and whey fraction, and skim milk provoked substantial immune responses at various degrees, indicating their potent immunogenicity. The protein mixtures prepared using the fractionated whey proteins with or without caseins appeared less immunogenic in both native and heat-treated forms, implying their potential of producing less immunogenic dairy products. The 100°C/30 s treatment significantly altered the immunogenicity of most of the potent protein stimulants, which mostly coincided with their levels of protein denaturation. The 72°C/15 s treatment caused the least protein denaturation but altered the immunogenicity of several protein stimulants notably, including heat-stable caseins and α-lactalbumin.

PENDUGAAN PEPTIDA BIOAKTIF DARI SUSU TERHIDROLISIS OLEH PROTEASE TUBUH DENGAN TEKNIK IN SILICO

The production of bioactive peptides catalyzed by gastrointestinal system (GIS) enzymes can be predicted in silico. The technique is more preferred than others such as in vivo and in vitro due to its low cost and less tedious procedure. The current study was aimed to predict bioactive peptides resulted from the digestion of bovine milk proteins. The digestion or so-called hydrolysis was simulated by means of a web-based in silico method. Identified bovine milk proteins from the available literatures were αS1-casein, αS2-casein, β-casein, κ-casein, β-lactoglobulin, α-lactalbumin, and lactoferrin. The compositions of amino acids (AAs) or protein sequences were accessed and tabulated from the Universal Protein Resource site (UniProt). Furthermore, the hydrolysis of each protein were simulated using three (3) GIS proteases, i.e., pepsin, trypsin, and chymotrypsin, and their possible combinations. All simulations were performed through web-based procedures using PeptideCutter, Expert Protein Analysis System (ExPASy). The resulted peptides were arranged according to the positions of cleavage sites for each cutting simulation, and compared to the available bioactive peptides data base in the literatures in terms of their AA residues (sequences). The simulation results indicated that β-casein and αS1-casein were the most potent proteins to yield bioactive peptides, of 52 and 48%, respectively. Moreover, each type of the investigated bovine milk proteins could be hydrolyzed by GIS proteases to produce antihypertensive bioactive peptides. This web-based in silico method is conclusively useful to predict bioactive peptides derived from bovine milk, and may also be used for other protein sources.

Therapeutics and Immunoprophylaxis Against Noroviruses and Rotaviruses: The Past, Present, and Future.

Background: Noroviruses and rotaviruses are important viral etiologies of severe gastroenteritis. Noroviruses are the primary cause of nonbacterial diarrheal outbreaks in humans, whilst rotaviruses are a major cause of childhood diarrhea. Although both enteric pathogens substantially impact human health and economies, there are no approved drugs against noroviruses and rotaviruses so far. On the other hand, whilst the currently licensed rotavirus vaccines have been successfully implemented in over 100 countries, the most advanced norovirus vaccine has recently completed phase-I and II trials.Methods: We performed a structured search of bibliographic databases for peer-reviewed research litera-ture on advances in the fields of norovirus and rotavirus therapeutics and immunoprophylaxis.Results: Technological advances coupled with a proper understanding of viral morphology and replication over the past decade has facilitated pioneering research on therapeutics and immunoprophylaxis against noroviruses and rotaviruses, with promising outcomes in human clinical trials of some of the drugs and vaccines. This review focuses on the various developments in the fields of norovirus and rotavirus thera-peutics and immunoprophylaxis, such as potential antiviral drug molecules, passive immunotherapies (oral human immunoglobulins, egg yolk and bovine colostral antibodies, llama-derived nanobodies, and anti-bodies expressed in probiotics, plants, rice grains and insect larvae), immune system modulators, probiot-ics, phytochemicals and other biological substances such as bovine milk proteins, therapeutic nanoparti-cles, hydrogels and viscogens, conventional viral vaccines (live and inactivated whole virus vaccines), and genetically engineered viral vaccines (reassortant viral particles, virus-like particles (VLPs) and other sub-unit recombinant vaccines including multi-valent viral vaccines, edible plant vaccines, and encapsulated viral particles).Conclusions: This review provides important insights into the various approaches to therapeutics and im-munoprophylaxis against noroviruses and rotaviruses..