Types Of Proteases Research Articles

Structure-based molecular insights into matrix metalloproteinase inhibitors in cancer treatments.

Protease inhibitors are of considerable interestas anticancer agents. Matrix metalloproteinases (MMPs) were the earliest type of proteases considered as anticancer targets. The developments of MMP inhibitors (MMPIs) by pharmaceutical companies can be dated from the early 1980s. Thus far, none of the over 50 MMPIs entering clinical trials have been approved. This work summarizes the reported studies on the structureof MMPs and complexes with ligands and inhibitors, based on which, the authorsanalyzed the clinical failures of MMPIs in a structural biological manner. Furthermore, MMPs were systematically comparedwith urokinase, a protease-generating plasmin, which plays similar pathological roles in cancer development;the reasons for the clinical successes of urokinase inhibitors andthe clinical failuresof MMPIs arediscussed.

The effectiveness of Proteolytic bacteria in the leather and detergent industry isolated waste from the Modjo tannery

Protease also called proteinase or peptidase is a digestive enzyme that is categorized under proteolytic enzymes and it has great potential in industrial application. Extracellular proteases are used in a variety of industries because they exhibit practically all of the characteristics needed for biotech applications such as detergent, bioremediation, food, and leather processing. In the synthesis of all three major types of acidic, neutral, and alkaline proteases, microbial sources have dominated an unbeatable area. Alkaline proteases are a large group of industrial enzymes formed by a wide variety of species, including animals, fungi, and bacteria. The fermentation method serves to make bacteria, fungi, and yeast alkaline proteases. Proteases are produced in large quantities by Gram-positive bacteria, especially those belonging to the Bacillus genus. Following standard procedures, the bacterial isolates PMOJ-01 and PMOJ-05 with the prominent zone of clearance and efficient enzyme development were further characterized to the genus level. Moreover, the growth conditions for the highest protease production were optimized with different pH, temperatures, and NaCl concentrations, in the results of PMOJ-01 and PMOJ- 05 pH (7 and 8), temperatures 45oC, and 1% NaCl concentrations both cases respectively. The proteases activities from PMOJ-01, Pseudomonas aeruginosa, and PMOJ-05, Bacillus subtilis were most active at pH 7.0 and pH 8.0 and temperature at 35oC and 45oC, respectively. The enzyme activity and the total solid protease sample of the crude enzyme of Pseudomonas aeruginosa and Bacillus subtilis were 0.299 U/ mL and 0.289 U/ mL, 1.37±0.14 U/mg, and 1.199 U/mg respectively. The effect on dehairing, distaining, and scum removal revealed that the purified protease enzyme of PMOJ-01 and PMOJ-05 can be used in detergent and leather industries.

The Angiostrongylus vasorum Excretory/Secretory and Surface Proteome Contains Putative Modulators of the Host Coagulation.

Angiostrongylus vasorum is a cardiopulmonary nematode of canids and is, among others, associated with bleeding disorders in dogs. The pathogenesis of such coagulopathies remains unclear. A deep proteomic characterization of sex specific A. vasorum excretory/secretory proteins (ESP) and of cuticular surface proteins was performed, and the effect of ESP on host coagulation and fibrinolysis was evaluated in vitro. Proteins were quantified by liquid chromatography coupled to mass spectrometry and functionally characterized through gene ontology and pathway enrichment analysis. In total, 1069 ESP (944 from female and 959 from male specimens) and 1195 surface proteins (705 and 1135, respectively) were identified. Among these were putative modulators of host coagulation, e.g., von Willebrand factor type D domain protein orthologues as well as several proteases, including serine type proteases, protease inhibitors and proteasome subunits. The effect of ESP on dog coagulation and fibrinolysis was evaluated on canine endothelial cells and by rotational thromboelastometry (ROTEM). After stimulation with ESP, tissue factor and serpin E1 transcript expression increased. ROTEM revealed minimal interaction of ESP with dog blood and ESP did not influence the onset of fibrinolysis, leading to the conclusion that Angiostrongylus vasorum ESP and surface proteins are not solely responsible for bleeding in dogs and that the interaction with the host’s vascular hemostasis is limited. It is likely that coagulopathies in A. vasorum infected dogs are the result of a multifactorial response of the host to this parasitic infection.

Effects of protease supplementation and diet type on jejunal and ileal digestibility and total tract metabolisability of nitrogen, starch, and energy in broilers

ABSTRACT 1. A study was conducted to assess the effects of supplemental protease (0 or 15,000 units/kg) and diet type (maize- or wheat-based) on apparent jejunal and ileal digestibility and apparent total tract metabolisability of nutrients in Cobb 500 mixed-sex broilers from 6 to 31 d of age. 2. Birds were randomly distributed into 56 metabolism cages (6 birds/cage; 14 replicates/treatment). At 22 d of age, jejunal and ileal digesta contents were collected and pooled from 4 birds/cage to determine apparent digestibility coefficients (DC) and digestible energy (DE). Feed intake was measured, and total excreta were collected from 8 to 11 and 18 to 21 d of age to determine apparent metabolisable energy (AME) and total tract nitrogen and starch metabolisability coefficients. 3. Broilers offered the maize-based diet with protease had greater (P < 0.05) jejunal nitrogen DC, starch DC, and DE (8.2, 6.5, and 14.9%, respectively) and ileal nitrogen DC and DE (5.1 and 6.8%, respectively) than those offered the maize-based diet without protease. Ileal starch DC was increased (P < 0.05) by 1.1% with protease supplementation. 4. Broilers offered maize-based diets had greater (P < 0.05) nitrogen (7.3%) and starch (0.6%) metabolisability coefficients and AME (4.7%) from 8 to 11 d of age, and nitrogen (4.3%) metabolisability coefficients and AME (2.0%) from 18 to 21 d of age compared with those offered wheat-based diets. 5. Thus, protease supplementation and diet type can affect digestive dynamics and nutrient utilisation in broilers.

Supplemental protease with phytase and xylanase and cereal grain source affected nutrient digestibility and performance of broilers

A study was conducted to evaluate the effects of supplemental protease and cereal grain type on nutrient digestibility (jejunum and ileum) and performance of broilers offered diets with reduced amino acid concentrations and supplemental xylanase and phytase. A total of 624 male broiler chicks (Ross 308) were randomly distributed into 48 floor pens (13 chicks/pen; 0.07 m2/bird) and offered one of six dietary treatments with eight replicates per treatment. Dietary treatments were either maize- or wheat-based with a positive control (PC) reference diet, a negative control diet without protease (NC; 60 g/kg lower amino acid density than PC), and an NC diet with protease. The reduction in amino acid density affected (P&lt;0.05) nutrient digestibility by varying degrees depending on cereal grain source. At 14 d of age, cereal grain and protease showed a significant interaction (P&lt;0.05) which affected jejunal and ileal starch digestibility, whereby protease increased digestibility in birds fed wheat-based diets but not in those fed maize-based diets. Cereal grain source affected (P&lt;0.05) nitrogen (jejunum and ileum) and digestible energy (DE; ileum), where birds fed wheat-based diets had higher digestibility than those fed maize-based diets. At 28 d of age, birds fed wheat-based diets had a higher (P&lt;0.01) jejunal and ileal nitrogen digestibility, whereas protease reduced ileal nitrogen digestion. Protease affected ileal starch digestion in birds fed wheat, but not maize-based diets, resulting in a significant cereal grain × protease interaction (P&lt;0.05). Wheat-based diets had a higher DE than maize-based diets in both the jejunum and ileum. From 15 to 35 d of age, cereal grain source (P&lt;0.05) affected performance, whereby broilers offered maize-based diets had better performance than those fed wheat-based diets.

Cloning, Characterization, Expression Analysis, and Agglutination Studies of Novel Gene Encoding β-d-Galactose, N-Acetyl-d-Glucosamine and Lactose-Binding Lectin from Rice Bean (Vigna umbellata)

Lectins are glycoproteins and known for their peculiar carbohydrate-binding activity and their insect-pest-resistant properties. Earlier we have published our research finding on novel gene encoding Bowman-Birk type protease inhibitor with insecticidal properties from rice bean. This paper presents first report on cloning, sequencing, and expression of RbL ORF of 843bp encoding 280 amino acids long lectin precursor from rice bean (Vigna umbellata) seeds. Blast analysis revealed more than 90% similarity of RbL protein with Vigna aconitifolia and Vigna angularis lectins. Phylogenetic analysis also revealed a close relationship between RbL and other legume lectins. Sequence analysis of genomic DNA revealed intronless nature of RbL gene (GenBank accession No. MT043160). The isolated RbL ORF was expressed in E. coli BL-21(DE3) cells and maximum expression was recorded with 0.5mM IPTG after 4h incubation at 37°C. Western blotting confirmed RbL protein expression in E. coli. Recombinant protein (His6-RbL) of ~ 35kDam.wt was purified using Ni-NTA affinity chromatography to the extent of 0.26mg/ml. In silico analysis characterized RbL protein as acidic, stable, hydrophobic, and secretary protein with one signal peptide cleavage site (A26-A27) and four N-glycosylation sites. Template-based 3D model of RbL was structured using MODELLER tool and validated as good quality model. Structural analysis revealed dominance of β-pleated sheets and β-turns in RbL protein structure. β-D-galactose, N-acetyl-D-glucosamine, and lactose were predicted as putative ligands for RbL protein. Hydrogen bonding and hydrophobic forces were the major interactions between the predicted ligands and RbL protein. Agglutination and agglutination inhibition assays confirmed the binding specificity of RbL protein with the trypsinized rabbit erythrocytes and with the predicted ligands, respectively. Gene ontology analysis functionally annotated RbL protein as a plant defense protein. The novel information generated in the study is not mere pre-experimental findings but could also lay foundation for future research on exploring RbL gene and encoding protein for different biomedical and biotechnological applications.

Formation of whey protein aggregates by partial hydrolysis and reduced thermal treatment

Thermal treatment to form functional, soluble whey protein aggregates often generates undesired off-flavours. The aim of the present study was to generate soluble aggregates of whey protein isolates (WPI) at lower temperature than traditionally used by introducing partial hydrolysis of WPI prior to thermal treatment. WPI was partially hydrolyzed either by Bacillus licheniformis protease (BLP) or by a trypsin type protease (TP) at pH 7.0 and 65 °C for 1–10 min prior to thermal treatment at 80 °C for 5–10 min to generate soluble protein aggregates. Partial hydrolysis by BLP and TP induced structural changes and exposure of free thiol groups, but no change of surface hydrophobicity of WPI. After incubation with BLP or TP and thermal treatment, the yield of protein aggregates was 36–48%, which was similar to the reference sample obtained by thermal treatment at 90 °C for 10 min. In addition to non-covalent interactions, disulfide bonds also contributed to the association of protein aggregates. The soluble protein aggregates obtained by BLP partial hydrolysis and thermal treatment, had particle size range of 10–100 nm in radius which was in the same range as observed for the reference sample; and samples obtained by TP treatment had a particle size range of 7–50 nm in radius. However, aggregates obtained by BLP or TP and thermal treatment were not UHT (140 °C/5 s) stable.

COVID-19 and features of cardiovascular involvement

The article provides an overview of current information on the pathogenesis of COVID-19 and organ-specific lesions developing in this disease. The data on inflammation and its biochemical markers, on the features of coagulopathy, endothelial damage and microthrombosis are presented in detail. Particular attention is paid to the role of receptors for angiotensin converting enzyme type 2 and transmembrane serine protease type 2 in the development of organ-specific lesions in COVID-19. The pathogenesis of damage to the cardiovascular system is considered in detail with the presentation of data from foreign literature on changes in the myocardium and the author's results of transthoracic echocardiographic examination in patients who have undergone COVID-19.

Dabrafenib, idelalisib and nintedanib act as significant allosteric modulator for dengue NS3 protease.

Dengue virus (DENV) encodes a unique protease (NS3/NS2B) essential for its maturation and infectivity and, it has become a key target for anti-viral drug design to treat dengue and other flavivirus related infections. Present investigation established that some of the drug molecules currently used mainly in cancer treatment are susceptible to bind non-active site (allosteric site/ cavity) of the NS3 protease enzyme of dengue virus. Computational screening and molecular docking analysis found that dabrafenib, idelalisib and nintedanib can bind at the allosteric site of the enzyme. The binding of the molecules to the allosteric site found to be stabilized via pi-cation and hydrophobic interactions, hydrogen-bond formation and π-stacking interaction with the molecules. Several interacting residues of the enzyme were common in all the five serotypes. However, the interaction/stabilizing forces were not uniformly distributed; the π-stacking was dominated with DENV3 proteases, whereas, a charged/ionic interaction was the major force behind interaction with DENV2 type proteases. In the allosteric cavity of protease from DENV1, the residues Lys73, Lys74, Thr118, Glu120, Val123, Asn152 and Ala164 were involved in active interaction with the three molecules (dabrafenib, idelalisib and nintedanib). Molecular dynamics (MD) analysis further revealed that the molecules on binding to NS3 protease caused significant changes in structural fluctuation and gained enhanced stability. Most importantly, the binding of the molecules effectively perturbed the protein conformation. These changes in the protein conformation and dynamics could generate allosteric modulation and thus may attenuate/alter the NS3 protease functionality and mobility at the active site. Experimental studies may strengthen the notion whether the binding reduce/enhance the catalytic activity of the enzyme, however, it is beyond the scope of this study.

Microbial proteases: ubiquitous enzymes with innumerable uses.

Proteases are ubiquitous enzymes, having significant physiological roles in both synthesis and degradation. The use of microbial proteases in food fermentation is an age-old process, which is today being successfully employed in other industries with the advent of ‘omics’ era and innovations in genetic and protein engineering approaches. Proteases have found application in industries besides food, like leather, textiles, detergent, waste management, agriculture, animal husbandry, cosmetics, and pharmaceutics. With the rising demands and applications, researchers are exploring various approaches to discover, redesign, or artificially synthesize enzymes with better applicability in the industrial processes. These enzymes offer a sustainable and environmentally safer option, besides possessing economic and commercial value. Various bacterial and fungal proteases are already holding a commercially pivotal role in the industry. The current review summarizes the characteristics and types of proteases, microbial source, their current and prospective applications in various industries, and future challenges. Promoting these biocatalysts will prove significant in betterment of the modern world.

Physicochemical properties and antioxidant activity of enzymatic hydrolysed chickpea (Cicer arietinum L.) protein as influence by alcalase and papain enzyme

Alcalase is a protease which prepared from the Bacillus licheniformis bacteria, whereas papain is a cysteine protease which prepared from plant sources papaya fruit latex, and both are utilized in enzymatic hydrolysis. However, each enzyme showed different effect during the hydrolysis process. In this present research, the effects of two commercial proteases, alcalase and papain, on the preparation and characterisation of physiochemical properties and antioxidant activity of chickpea (Cicer arietinum L.) protein hydrolysate (CPH) were studied. Different parameters were used during hydrolysis using alcalase (pH 10, temperature 45 °C) and papain (pH 6.7, temperature 65 °C). CPH hydrolysed by enzyme alcalase (CPHA) showed higher yields (32.5%) and degree of hydrolysis (DH) (8.62%) than CPH hydrolysed by enzyme papain (CPHP), which had lower yields (13.92%) and DH (7.99%). In addition, CPHA led to a lower molecular weight (10 kDa) than CPHP (14.3 kDa). However, in terms of chemical composition and solubility, no significant differences existed between CPHA and CPHP. Moreover, CPHA showed significantly higher foaming capacity, foaming stability, emulsifying stability, antioxidant activity on DPPH radical scavenging activity as compared to CPHP. CPHA also showed significantly lower emulsifying capacity than CPHP. These findings indicate that the type of protease used to hydrolyse CPH has a significant impact on the physicochemical properties and antioxidant activity of the produced CPH. • The yield of CPHA was higher (32.5%) than CPHP (13.92%). • CPHA showed a higher DH value however lower in molecular weight value than CPHP. • CPHA had a better foaming capacity, stability, and emulsifying stability than CPHP. • CPHA showed a lower emulsifying capacity (52.94%) than CPHP (57.33%). • CPHA had higher antioxidant activity in DPPH radical scavenging activity (79.57%).

An Updated review on production of food derived bioactive peptides; focus on the psychrotrophic bacterial proteases

Psychrotrophs or psychrophiles are extremophilic microorganisms that have the maximum temperature above 20 °C for growth. Since around 70% of the earth's biosphere temperature is below 5 °C, psychrotrophs could be found in diversity of species. These bacteria are source of enzymes, mainly including proteases used in different food industries where main reactions should be proceeded in low temperatures to save the aroma and flavor of food materials and bioactivity of valuable food ingredients such as bioactive peptides. Proteases are among the major groups of industrial enzymes accounting for 60% of the total global trade of enzymes. In this review, at first, we describe the biology, classification and physiological functions of different proteases obtained from psychrotrophic bacteria. Then, we present the various methods to produce and purify these enzymes from the bacteria. Moreover, we discuss several applications of the proteases to produce bioactive peptides from versatile protein sources. A comprehensive introduction to different types of proteases and their production and purification methods could help researchers identify novel proteases with improved properties, considering that their total market is significantly growing every year. • Psychrotrophic bacteria are one of the main sources of enzymes production. • Proteases are one of the major groups of industrial enzymes in global trade. • Herein a classifications and application of psychrotrophic proteases is presented. • Researchers who are seeking novel applicable proteases can use this review.

Augmenting Feeding Value of Rice Distillers Dried Grain with Solubles Through Dietary Addition of Enzymes in Broilers

Background: Substitution of expensive protein sources like soybean meal with lower cost ingredient like rice distillers dried grain with solubles (rDDGS) would reduce the cost of the feed. Various enzymes supplementation may be used to increase their inclusion level. Methods: An in-vivo study of 42 days was undertaken (n= 384) in broilers divided into 12 dietary treatments as per 3x4 factorial design. Two levels of rDDGS consisted of 12.5 and 15% without and with three different types of enzymes xylanase, protease and multienzymes were used to find out substract specific enzyme. Result: Overall the growth performance of broiler chicken was significantly lower at 15% rDDGS level as compared to control and 12.5% rDDGS level. However, it was significantly (P less than 0.01) improved by supplementation of any of the three enzymes (xylanase, protease and multienzymes) in 12.5 and 15% rDDGS groups. Nitrogen retention, dry matter and energy metabolizability were significantly (P less than 0.01) improved in all enzymes supplemented groups. The cost of feed per kg live weight and meat yield was significantly (P less than 0.01) lower at 12.5% rDDGS with enzymes supplementation. Thus, it is concluded that the enzymes supplementation can increase inclusion level of rDDGS from 12.5 to 15% for economic broiler production.

Development of a Bio-Layer Interferometry-Based Protease Assay Using HIV-1 Protease as a Model

Proteolytic enzymes have great significance in medicine and the pharmaceutical industry and are applied in multiple fields of life sciences. Therefore, cost-efficient, reliable and sensitive real-time monitoring methods are highly desirable to measure protease activity. In this paper, we describe the development of a new experimental approach for investigation of proteolytic enzymes. The method was designed by the combination of recombinant fusion protein substrates and bio-layer interferometry (BLI). The protease (PR) of human immunodeficiency virus type 1 (HIV-1) was applied as model enzyme to set up and test the method. The principle of the assay is that the recombinant protein substrates immobilized to the surface of biosensor are specifically cleaved by the PR, and the substrate processing can be followed by measuring change in the layer thickness by optical measurement. We successfully used this method to detect the HIV-1 PR activity in real time, and the initial rate of the signal decrease was found to be proportional to the enzyme activity. Substrates representing wild-type and modified cleavage sites were designed to study HIV-1 PR’s specificity, and the BLI-based measurements showed differential cleavage efficiency of the substrates, which was proven by enzyme kinetic measurements. We applied this BLI-based assay to experimentally confirm the existence of extended binding sites at the surface of HIV-1 PR. We found the measurements may be performed using lysates of cells expressing the fusion protein, without primary purification of the substrate. The designed BLI-based protease assay is high-throughput-compatible and enables real-time and small-volume measurements, thus providing a new and versatile approach to study proteolytic enzymes.

Broad Influence of Mutant Ataxin-3 on the Proteome of the Adult Brain, Young Neurons, and Axons Reveals Central Molecular Processes and Biomarkers in SCA3/MJD Using Knock-In Mouse Model.

Spinocerebellar ataxia type 3 (SCA3/MJD) is caused by CAG expansion mutation resulting in a long polyQ domain in mutant ataxin-3. The mutant protein is a special type of protease, deubiquitinase, which may indicate its prominent impact on the regulation of cellular proteins levels and activity. Yet, the global model picture of SCA3 disease progression on the protein level, molecular pathways in the brain, and neurons, is largely unknown. Here, we investigated the molecular SCA3 mechanism using an interdisciplinary research paradigm combining behavioral and molecular aspects of SCA3 in the knock-in ki91 model. We used the behavior, brain magnetic resonance imaging (MRI) and brain tissue examination to correlate the disease stages with brain proteomics, precise axonal proteomics, neuronal energy recordings, and labeling of vesicles. We have demonstrated that altered metabolic and mitochondrial proteins in the brain and the lack of weight gain in Ki91 SCA3/MJD mice is reflected by the failure of energy metabolism recorded in neonatal SCA3 cerebellar neurons. We have determined that further, during disease progression, proteins responsible for metabolism, cytoskeletal architecture, vesicular, and axonal transport are disturbed, revealing axons as one of the essential cell compartments in SCA3 pathogenesis. Therefore we focus on SCA3 pathogenesis in axonal and somatodendritic compartments revealing highly increased axonal localization of protein synthesis machinery, including ribosomes, translation factors, and RNA binding proteins, while the level of proteins responsible for cellular transport and mitochondria was decreased. We demonstrate the accumulation of axonal vesicles in neonatal SCA3 cerebellar neurons and increased phosphorylation of SMI-312 positive adult cerebellar axons, which indicate axonal dysfunction in SCA3. In summary, the SCA3 disease mechanism is based on the broad influence of mutant ataxin-3 on the neuronal proteome. Processes central in our SCA3 model include disturbed localization of proteins between axonal and somatodendritic compartment, early neuronal energy deficit, altered neuronal cytoskeletal structure, an overabundance of various components of protein synthesis machinery in axons.

PINIR: a comprehensive information resource for Pin-II type protease inhibitors

BackgroundSerine protease inhibitors belonging to the Potato type-II Inhibitor family Protease Inhibitors (Pin-II type PIs) are essential plant defense molecules. They are characterized by multiple inhibitory repeat domains, conserved disulfide bond pattern, and a tripeptide reactive center loop. These features of Pin-II type PIs make them potential molecules for protein engineering and designing inhibitors for agricultural and therapeutic applications. However, the diversity in these PIs remains unexplored due to the lack of annotated protein sequences and their functional attributes in the available databases.ResultsWe have developed a database, PINIR (Pin-II type PIs Information Resource), by systematic collection and manual annotation of 415 Pin-II type PI protein sequences. For each PI, the number and position for signature sequences are specified: 695 domains, 75 linkers, 63 reactive center loops, and 10 disulfide bond patterns are identified and mapped. Database analysis revealed novel subcategories of PIs, species-correlated occurrence of inhibitory domains, reactive center loops, and disulfide bond patterns. By analyzing linker regions, we predict that alternative processing at linker regions could generate PI variants in the Solanaceae family.ConclusionPINIR (https://pinir.ncl.res.in) provides a web interface for browsing and analyzing the protein sequences of Pin-II type PIs. Information about signature sequences, spatio-temporal expression, biochemical properties, gene sequences, and literature references are provided. Analysis of PINIR depicts conserved species-specific features of Pin-II type PI protein sequences. Diversity in the sequence of inhibitory domains and reactive loops directs potential applications to engineer Pin-II type PIs. The PINIR database will serve as a comprehensive information resource for further research into Pin-II type PIs.

Technological and bioactive properties of wheat glutenin hydrolysates prepared with various commercial proteases

This study aimed to investigate the effect of different types of proteolytic enzymes (flavourenzyme, savinase, subsitilin, and savinase) on the technological and bioactive properties of glutenin hydrolysate. The degree of hydrolysis (DH) of the glutenin proteins significantly differed according to the enzyme types (p < 0.05). SDS patterns showed that the number and density of the bands decreased as the DH increased and the lowest molecular weight peptide fractions was observed in savinase and subsitilin hydrolysates. In general, hydrolysates with low DH had superior technological properties (oil absorption capacity, hydrophobicity, emulsifying activity, and stability index) compared to the higher ones. After enzymatic hydrolysis, the β sheet (%) of the hydrolysates increased, and the β turn (%) decreased compared to the control sample. The antioxidant capacity of the glutenin significantly increased by enzymatic hydrolysis (p < 0.05). The highest ABTS radical scavenging activity and FRAP activity was found in savinase and subsitilin hydrolysates with relatively higher DH. All hydrolysates exhibited an anti-diabetic effect depending on the DH. The anti-diabetic activity of the hydrolysate increased with increasing DH value. Hydrolysate samples showed no cytotoxic effect on the HEK293 healthy cell line, whereas hydrolysates exhibited cytotoxic effects on MIA PaCa 2 and CaCo 2 cancer cell lines.

Cost Economics of Feeding Rice Gluten Meal and Soybean Meal through Dietary Addition of Enzymes in Broilers

Background: The rice gluten meal a relatively newer feedstuff is by-product of wet-milling of rice obtained after starch extraction and syrup preparation. The rice gluten meal is high crude protein and energy ingredient which is priced lower than soybean meal. Substitution of expensive protein sources with lower cost ingredients would potentially reduce the cost of the feed.Methods: A biological experiment of 42 days duration was undertaken in day old chicks (n= 384) divided into 12 dietary treatments as per 3x4 factorial design having 4 replicates per treatment with 8 birds in each. Twelve experimental diets were prepared by incorporating control, two different levels of rice gluten meal RGM consisted of (15 and 17.5%), without and with three different types of enzymes xylanase (X), protease (P) and multienzymes (M).Result: The rice gluten meal at the inclusion level of 15% reduced feed cost per kg live weight, meat yield and eviscerated yield by 5.44, 5.13 and 4.69%, respectively. Protease enzyme supplementation in 15% rice gluten meal further reduced feed cost per kg live weight by 2.60%. Thus, it may be concluded protease enzyme is most cost effective in rice gluten meal diet.

Effect of Protease Type and Peptide Size on the In Vitro Antioxidant, Antihypertensive and Anti-Diabetic Activities of Eggplant Leaf Protein Hydrolysates.

Solanum macrocarpon (eggplant) leaf protein isolate (ELI) was hydrolyzed using four different enzymes to produce hydrolysates from alcalase (AH), chymotrypsin (CH) pepsin (PH) and trypsin (TH). CH had an overall stronger antioxidant property and was separated using ultrafiltration membranes into <1, 1–3 and 3–5 kDa peptide fractions. Gel-permeation chromatography confirmed conversion of the ELI (average of 22 kDa) into protein hydrolysates that contained smaller peptides (<6 kDa). A total of 23 peptides consisting of tri and tetrapeptides were identified from the CH, which is a wider spectrum when compared to seven for AH and four each for TH and PH. CH exhibited stronger scavenging activities against DPPH and hydroxyl radicals. CH and TH exhibited the strongest inhibitions against angiotensin-converting enzyme. In contrast, AH was the strongest inhibitor of α-amylase while AH and PH had strong inhibitory activities against α-glucosidase when compared with other hydrolysates. Ultrafiltration fractionation produced peptides that were stronger (p < 0.05) scavengers of DPPH, and hydroxyl radicals, in addition to better metal-chelating and enzyme inhibition agents. The study concluded that the eggplant protein hydrolysates and the UF fractions may find applications in tackling oxidative stress-related diseases and conditions involving excessive activities of the metabolic enzymes.

Meat Waste Valorization through Protein Hydrolysis using Different Types of Proteases

Five different commercial proteases (Alcalase 2.4L, Flavourzyme 1000L, Neutrase, Protamex, and PTN) were evaluated for the simultaneous recovery of protein and lipids through hydrolysis. The hydrolysis reaction was monitored using the pH-stat procedure, in which samples were collected after 240 min of hydrolysis using each enzyme. The samples were analyzed for the degree of hydrolysis, protein hydrolysate, collagen, lipids, and fatty acids. A clear relationship was observed between the degree of hydrolysis and the amounts of recovered products. Serine endopeptidases from the microbial source (Alcalase) resulted in the maximum degree of hydrolysis (27.5%), lipid recovery (82.6%), and protein hydrolysates quality (average molecular weight of the hydrolysates = 472 Dalton), followed by formulations of serine protease and metalloprotease from the microbial source (Protamex). Metalloproteases from the microbial source (Neutrase) resulted in maximum collagen recovery (87.1%). Serine endopeptidases from the animal source (PTN) and endo/exopeptidases from the fungal source (Flavourzyme) exhibited an intermediate efficacy between Alcalase and Neutrase. In the case of all proteases, the product fatty acid profile matched well with that of the meat waste, which suggests that no chemical changes occurred in the lipids after the hydrolysis with the evaluated proteases. These results suggest that hydrolysis using proteases could serve as an ecofriendly and viable alternative for obtaining additional value from meat waste.