Potentials, prospects and applications of genome editing technologies in livestock production.

In recent years, significant progress has been achieved in genome editing applications using new programmable DNA nucleases such as zinc finger nucleases (ZFNs), transcription activator-like endonucleases (TALENs) and the clustered regularly interspaced short palindromic repeats/Cas9 system (CRISPR/Cas9). These genome editing tools are capable of nicking DNA precisely by targeting specific sequences, and enable the addition, removal or substitution of nucleotides via double-stranded breakage at specific genomic loci. CRISPR/Cas system, one of the most recent genome editing tools, affords the ability to efficiently generate multiple genomic nicks in single experiment. Moreover, CRISPR/Cas systems are relatively easy and cost effective when compared to other genome editing technologies. This is in part because CRISPR/Cas systems rely on RNA-DNA binding, unlike other genome editing tools that rely on protein-DNA interactions, which affords CRISPR/Cas systems higher flexibility and more fidelity. Genome editing tools have significantly contributed to different aspects of livestock production such as disease resistance, improved performance, alterations of milk composition, animal welfare and biomedicine. However, despite these contributions and future potential, genome editing technologies also have inherent risks, and therefore, ethics and social acceptance are crucial factors associated with implementation of these technologies. This review emphasizes the impact of genome editing technologies in development of livestock breeding and production in numerous species such as cattle, pigs, sheep and goats. This review also discusses the mechanisms behind genome editing technologies, their potential applications, risks and associated ethics that should be considered in the context of livestock.

Emergence, evolution, and vaccine production approaches of SARS-CoV-2 virus: Benefits of getting vaccinated and common questions.

The emergence of coronavirus disease 2019 (COVID-19) pandemic in Wuhan city, China at the end of 2019 made it urgent to identify the origin of the causal pathogen and its molecular evolution, to appropriately design an effective vaccine. This study analyzes the evolutionary background of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2 or SARS-2) in accordance with its close relative SARS-CoV (SARS-1), which was emerged in 2002. A comparative genomic and proteomic study was conducted on SARS-2, SARS-1, and Middle East respiratory syndrome coronavirus (MERS), which was emerged in 2012. In silico analysis inferred the genetic variability among the tested viruses. The SARS-1 genome harbored 11 genes encoding 12 proteins, while SARS-2 genome contained only 10 genes encoding for 10 proteins. MERS genome contained 11 genes encoding 11 proteins. The analysis also revealed a slight variation in the whole genome size of SARS-2 comparing to its siblings resulting from sequential insertions and deletions (indels) throughout the viral genome particularly ORF1AB, spike, ORF10 and ORF8. The effective indels were observed in the gene encoding the spike protein that is responsible for viral attachment to the angiotensin-converting enzyme 2 (ACE2) cell receptor and initiating infection. These indels are responsible for the newly emerging COVID-19 variants αCoV, ßCoV, γCoV and δCoV. Nowadays, few effective COVID-19 vaccines developed based on spike (S) glycoprotein were approved and become available worldwide. Currently available vaccines can relatively prevent the spread of COVID-19 and suppress the disease. The traditional (killed or attenuated virus vaccine and antibody-based vaccine) and innovated vaccine production technologies (RNA- and DNA-based vaccines and viral vectors) are summarized in this review. We finally highlight the most common questions related to COVID-19 disease and the benefits of getting vaccinated.

Green nanotechnology for controlling bacterial load and heavy metal accumulation in Nile tilapia fish using biological selenium nanoparticles biosynthesized by Bacillus subtilis AS12.

Heavy metal accumulation and pathogenic bacteria cause adverse effects on aquaculture. The active surface of selenium (Se) nanoparticles can mitigate these effects. The present study used Se-resistant Bacillus subtilis AS12 to fabricate biological Se nanoparticles (Bio-SeNPs). The double-edged Bio-SeNPs were tested for their ability to reduce the harmful effects of heavy metals and bacterial load in Nile tilapia (Oreochromis niloticus) and their respective influences on fish growth, behavior, and health. The Bio-SeNPs have a spherical shape with an average size of 77 nm and high flavonoids and phenolic content (0.7 and 1.9 g g-1 quercetin and gallic acid equivalents, respectively), resulting in considerable antioxidant and antibacterial activity. The Bio-SeNPs (3-5 µg ml-1) in the current study resolved two serious issues facing the aquaculture industry, firstly, the population of pathogenic bacteria, especially Aeromonas hydrophilia, which was reduced by 28-45% in fish organs. Secondly, heavy metals (Cd and Hg) at two levels (1 and 2 µg ml-1) were reduced by 50-87% and 57-73% in response to Bio-SeNPs (3-5 µg ml-1). Thus, liver function parameters were reduced, and inner immunity was enhanced. The application of Bio-SeNPs (3-5 µg ml-1) improved fish gut health, growth, and behavior, resulting in fish higher weight gain by 36-52% and a 40% specific growth rate, compared to controls. Furthermore, feeding and arousal times increased by 20-22% and 28-53%, respectively, while aggression time decreased by 78% compared to the control by the same treatment. In conclusion, Bio-SeNPs can mitigate the accumulation of heavy metals and reduce the bacterial load in a concentration-dependent manner, either in the fish media or fish organs.

Antibacterial Peptides Produced by Alcalase from Cowpea Seed Proteins.

Cowpea seed protein hydrolysates (CPH) were output from cowpea seeds applying alcalase® from Bacillus licheniformis. CPH with an elevated level of hydrolysis was fractionated by size exclusion chromatography (SEC). Both CPH and SEC-portions showed to contain antimicrobial peptides (AMPs) as they inhibited both Gram-positive bacteria, such as Listeria monocytogenes LMG10470 (L. monocytogenes), Listeria innocua. LMG11387 (L. innocua), Staphylococcus aureus ATCC25923 (S.aureus), and Streptococcus pyogenes ATCC19615 (St.pyogenes), and Gram-negative bacteria, such as Klebsiella pnemoniae ATCC43816 (K. pnemoniae), Pseudomonas aeroginosa ATCC26853 (P. aeroginosa), Escherichia coli ATCC25468) (E.coli) and Salmonella typhimurium ATCC14028 (S. typhimurium).The data exhibited that both CPH and size exclusion chromatography-fraction 1 (SEC-F1) showed high antibacterial efficiency versus almost all the assessed bacteria. The MIC of the AMPs within SEC-F1 and CPHs were (25 µg/mL) against P. aeruginosa, E.coli and St. pyogenes. However, higher MICsof approximately 100-150 µg/mL showed for both CPHs and SEC-F1 against both S. aureus and L. innocua; it was 50 µg/mL of CPH against S.aureus. The Electro-spray-ionization-mass-spectrometry (ESI-MS) of fraction (1) revealed 10 dipeptides with a molecular masses arranged from 184 Da to 364 Da and one Penta peptide with a molecular mass of approximately 659 Da inthe case of positive ions. While the negative ions showed 4 dipeptides with the molecular masses that arranged from 330 Da to 373 Da. Transmission electron microscope (TEM) demonstrated that the SEC-F1 induced changes in the bacterial cells affected. Thus, the results suggested that the hydrolysis of cowpea seed proteins by Alcalase is an uncomplicated appliance to intensify its antibacterial efficiency.

Biochemical and Functional Characterization of Kidney Bean Protein Alcalase-Hydrolysates and Their Preservative Action on Stored Chicken Meat.

A new preservation approach is presented in this article to prolong the lifetime of raw chicken meat and enhance its quality at 4 °C via coating with highly soluble kidney bean protein hydrolysate. The hydrolysates of the black, red, and white kidney protein (BKH, RKH, and WKH) were obtained after 30 min enzymatic hydrolysis with Alcalase (E/S ratio of 1:100, hydrolysis degree 25-29%). The different phaseolin subunits (8S) appeared in SDS-PAGE in 35-45 kD molecular weight range while vicilin appeared in the molecular weight range of 55-75 kD. The kidney bean protein hydrolysates have considerable antioxidant activity as evidenced by the DPPH-scavenging activity and ß-carotine-linolenic assay, as well as antimicrobial activity evaluated by disc diffusion assay. BKH followed by RKH (800 µg/mL) significantly (p ≤ 0.05) scavenged 95, 91% of DPPH and inhibited 82-88% of linoleic oxidation. The three studied hydrolysates significantly inhibited the growth of bacteria, yeast, and fungi, where BKH was the most performing. Kidney bean protein hydrolysates could shield the chicken meat because of their amphoteric nature and many functional properties (water and oil-absorbing capacity and foaming stability). The quality of chicken meat was assessed by tracing the fluctuations in the chemical parameters (pH, met-myoglobin, lipid oxidation, and TVBN), bacterial load (total bacterial count, and psychrophilic count), color parameters and sensorial traits during cold preservation (4 °C). The hydrolysates (800 µg/g) significantly p ≤ 0.05 reduced the increment in meat pH and TVBN values, inhibited 59-70% of lipid oxidation as compared to control during 30 days of cold storage via eliminating 50% of bacterial load and maintained secured storage for 30 days. RKH and WKH significantly (p ≤ 0.05) enhanced L*, a* values, thus augmented the meat whiteness and redness, while, BKH increased b* values, declining all color parameters during meat storage. RKH and WKH (800 µg/g) (p ≤ 0.05) maintained 50-71% and 69-75% of meat color and odor, respectively, increased the meat juiciness after 30 days of cold storage. BKH, RKH and WKH can be safely incorporated into novel foods.

Transfer of Anthocyanin Accumulating Delila and Rosea1 Genes from the Transgenic Tomato Micro-Tom Cultivar to Moneymaker Cultivar by Conventional Breeding.

Delila and Rosea1 anthocyanin accumulation genes were subjected to bioinformatics analysis. Delila protein has 56-69% similarity with different anthocyanin-rich plants, while Rosea1 protein has 83-87% with anthocyanin-rich plant proteins. This study aimed at transferring Delila and Rosea1 genes from the transgenic Micro-tom tomato cultivar to the Moneymaker tomato cultivar using traditional breeding for enhancing their fruit anthocyanin content. Results of all produced F1 plants of manual hybridization between both cultivars were consistent with the Mendelian inheritance hypothesis. Plants of F2 populations showed a 3:1 Mendelian segregation proportion (75% of plants have anthocyanin pigmentation). Seeds of F2 were individually cultured to get four homozygous lines with anthocyanin accumulation in fruits. The total anthocyanin in the anthocyanin-enriched inbred fruit (3 g/kg DM) represented a relative increase of about 131% of the parent level. The total phenolic compounds in inbred tomato fruits were 54.9 mg/100 g DM referring to a relative increase of about 51% of the respective parent plant. The antioxidant activity of inbred fruit at maturity (m) was 83.5% compared with 91% for TBHQ. The inbred (m) tomato fruit extract reduced the growth of G- bacteria G+ bacteria by 99% and 95%, respectively.

Aspergillus nidulans thermostable arginine deiminase-Dextran conjugates with enhanced molecular stability, proteolytic resistance, pharmacokinetic properties and anticancer activity.

The potential anticancer activity of arginine deiminase (ADI) via deimination of l-arginine into citrulline has been extensively verified against various arginine-auxotrophic tumors, however, the higher antigenicity, structural instability and in vivo proteolysis are the major challenges that limit this enzyme from further clinical implementation. Since, this clinically applied enzyme was derived from Mycobacterium spp, thus, searching for ADI from eukaryotic microbes "especially thermophilic fungi" could have a novel biochemical, conformational and catalytic properties. Aspergillus nidulans ADI was purified with 5.3 folds, with molecular subunit structure 48 kDa and entire molecular mass 120 kDa, ensuring its homotrimeric identity. The peptide fingerprinting analysis revealing the domain Glu95-Gly96-Gly97 as the conserved active site of A. nidulans ADI, with higher proximity to Mycobacterium ADI clade IV. In an endeavor to fortify the structural stability and anticancer activity of A. nidulans ADI, the enzyme was chemically modified with dextran. The optimal activity of Dextran-ADI conjugates was determined at 0.08:20 M ratio of ADI: Dextran, with an overall increase to ADI molecular subunit mass to ˜100 kDa. ADI was conjugated with dextran via the ε-amino groups interaction of surface lysine residues of ADI. The resistance of Dextran-ADI conjugate to proteolysis had been increased by 2.5 folds to proteinase K and trypsin, suggesting the shielding of >50% of ADI surface proteolytic recognition sites. The native and Dextran-ADI conjugates have the same optimum reaction temperature (37 °C), reaction pH and pH stability (7.0-8.0) with dependency on K+ ions as a cofactor. Dextran-ADI conjugates exhibited a higher thermal stability by ˜ 2 folds for all the tested temperatures, ensuring the acquired structural and catalytic stability upon dextran conjugation. Dextran conjugation slightly protect the reactive amino and thiols groups of surface amino acids of ADI from amino acids suicide inhibitors. The affinity of ADI was increased by 5.3 folds to free L-arginine with a dramatic reduction in citrullination of peptidylarginine residues upon dextran conjugation. The anticancer activity of ADI to breast (MCF-7), liver (HepG-2) and colon (HCT8, HT29, DLD1 and LS174 T) cancer cell lines was increased by 1.7 folds with dextran conjugation in vitro. Pharmacokinetically, the half-life time of ADI was increased by 1.7 folds upon dextran conjugation, in vivo. From the biochemical and hematological parameters, ADIs had no signs of toxicity to the experimental animals. In addition to the dramatic reduction of L-arginine in serum, citrulline level was increased by 2.5 folds upon dextran conjugation of ADI. This is first report exploring thermostable ADI from thermophilic A. nidulans with robust structural stability, catalytic efficiency and proteolytic resistance.

Restoring the Taxol biosynthetic machinery of Aspergillus terreus by Podocarpus gracilior Pilger microbiome, with retrieving the ribosome biogenesis proteins of WD40 superfamily.

Attenuating the Taxol yield of Aspergillus terreus with the subculturing and storage were the technical challenges that prevent this fungus to be a novel platform for industrial Taxol production. Thus, the objective of this study was to unravel the metabolic machineries of A. terreus associated with attenuation of Taxol productivity, and their restoring potency upon cocultivation with the Podocarpus gracilior microbiome. The Taxol yield of A. terreus was drastically reduced with the fungal subculturing. At the 10th subculture, the yield of Taxol was reduced by four folds (78.2 µg/l) comparing to the original culture (268 µg/l), as authenticated from silencing of molecular expression of the Taxol-rate limiting enzymes (GGPPS, TDS, DBAT and BAPT) by qPCR analyses. The visual fading of A. terreus conidial pigmentation with the subculturing, revealing the biosynthetic correlation of melanin and Taxol. The level of intracellular acetyl-CoA influx was reduced sequentially with the fungal subculturing, rationalizing the decreasing on Taxol and melanin yields. Fascinatingly, the Taxol biosynthetic machinery and cellular acetyl-CoA of A. terreus have been completely restored upon addition of 3% surface sterilized leaves of P. gracilior, suggesting the implantation of plant microbiome on re-triggering the molecular machinery of Taxol biosynthesis, their transcriptional factors, and/or increasing the influx of Acetyl-CoA. The expression of the proteins of 74.4, 68.2, 37.1 kDa were exponentially suppressed with A. terreus subculturing, and strongly restored upon addition of P. gracilior leaves, ensuring their profoundly correlation with the molecular expression of Taxol biosynthetic genes. From the proteomic analysis, the restored proteins 74.4 kDa of A. terreus upon addition of P. gracilior leaves were annotated as ribosome biogenesis proteins YTM and microtubule-assembly proteins that belong to WD40 superfamily. Thus, further ongoing studies for molecular cloning and expression of these genes with strong promotors in A. terreus, have been initiated, to construct a novel platform of metabolically stable A. terreus for sustainable Taxol production. Attenuating the Taxol yield of A. terreus with the multiple-culturing and storage might be due to the reduction on main influx of acetyl-CoA, or downregulation of ribosome biogenesis proteins that belong to WD40 protein superfamily.

Purification and Characterization of Ornithine Decarboxylase from Aspergillus terreus; Kinetics of Inhibition by Various Inhibitors.

l-Ornithine decarboxylase (ODC) is the rate-limiting enzyme of de novo polyamine synthesis in humans and fungi. Elevated levels of polyamine by over-induction of ODC activity in response to tumor-promoting factors has been frequently reported. Since ODC from fungi and human have the same molecular properties and regulatory mechanisms, thus, fungal ODC has been used as model enzyme in the preliminary studies. Thus, the aim of this work was to purify ODC from fungi, and assess its kinetics of inhibition towards various compounds. Forty fungal isolates were screened for ODC production, twenty fungal isolates have the higher potency to grow on L-ornithine as sole nitrogen source. Aspergillus terreus was the most potent ODC producer (2.1 µmol/mg/min), followed by Penicillium crustosum and Fusarium fujikuori. These isolates were molecularly identified based on their ITS sequences, which have been deposited in the NCBI database under accession numbers MH156195, MH155304 and MH152411, respectively. ODC was purified and characterized from A. terreus using SDS-PAGE, showing a whole molecule mass of ~110 kDa and a 50 kDa subunit structure revealing its homodimeric identity. The enzyme had a maximum activity at 37 °C, pH 7.4-7.8 and thermal stability for 20 h at 37 °C, and 90 days storage stability at 4 °C. A. terreus ODC had a maximum affinity (Km) for l-ornithine, l-lysine and l-arginine (0.95, 1.34 and 1.4 mM) and catalytic efficiency (kcat/Km) (4.6, 2.83, 2.46 × 10-5 mM-1·s-1). The enzyme activity was strongly inhibited by DFMO (0.02 µg/mL), curcumin (IC50 0.04 µg/mL), propargylglycine (20.9 µg/mL) and hydroxylamine (32.9 µg/mL). These results emphasize the strong inhibitory effect of curcumin on ODC activity and subsequent polyamine synthesis. Further molecular dynamic studies to elucidate the mechanistics of ODC inhibition by curcumin are ongoing.

Effect of a 24-week weight management program on serum leptin level in correlation to anthropometric measures in obese female: A randomized controlled clinical trial.

BACKGROUND: Obesity is a major contributor to preventable disease and death across the globe. Obesity is complex. Although its risk factors are myriad and compounding, there is an urgent need for a deeper understanding of the way risk factors interact with each other. Leptin is a peptide regulates food intake and body weight. However, the notion of leptin as an anti-obesity hormone was called into question because obesity is typically associated with high leptin levels and not leptin deficiency thus, we aimed to measure leptin levels in obese female in correlation to anthropometric measures and to evaluate the impact of weight loss on its level and metabolic parameters. SUBJECT AND METHODS: case-control study enrolled 40 control groups, 50 obese women. We measured anthropometric measures BMI, Waist/hip ratio (WHR). Fat mass index (FMI%) and free fat mass index (FFMI%) were assessed by dual energy X-Ray absorptiometry (DEXA) The serum levels of leptin were measured by ELISA. RESULTS: Our results revealed that serum leptin levels were higher in obese women compared to controls. Moreover, it was positively correlated to anthropometric measures, glycemic and lipid profile. Linear regression analysis revealed that BMI was the main independent studied parameters associated with serum leptin level among other clinical and laboratory biomarkers. Interestingly, after 12 weeks of following the Mediterranean diet (MD)-based weight loss program, serum leptin levels were decreased. Logistic regression analysis was performed to detect the main predictors' biomarkers associated with weight loss among obese women. We found that serum leptin and FMI% were an independent predictor of response with odds ratios of 1.69 and 1.64 respectively (P < 0.001), Receiver operating characteristic analyses revealed that the AUC of serum leptin in discriminating obese women from lean ones was 0.893 (95% CI = 0.815-0.917) with sensitivity = 90%, specificity = 96%, and the cutoff values was 36.32 ng/ml. CONCLUSION: Serum leptin could be a valuable diagnostic marker of obesity and its comorbidities. Moreover, significant weight loss leads to decrease serum leptin levels and improvement of glycemic and lipid profiles.

Biochemical characterization of peptidylarginine deiminase-like orthologs from thermotolerant Emericella dentata and Aspergillus nidulans.

Peptidylarginine deiminases (PADs) are a group of hydrolases, mediating the deimination of peptidylarginine residues into peptidyl-citrulline. Equivocal protein citrullination by PADs of fungal pathogens has a strong relation to the progression of multiple human diseases, however, the biochemical properties of fungal PADs remain ambiguous. Thus, this is the first report exploring the molecular properties of PAD from thermotolerant fungi, to imitate the human temperature. The teleomorph Emericella dentata and anamorph Aspergillus nidulans have been morphologically and molecularly identified, with observed robust growth at 37-40 °C, and strong PAD productivity. The physiological profiles of E. dentata and A. nidulans for PADs production in response to carbon, nitrogen sources, initial medium pH and incubation temperature were relatively identical, emphasizing the taxonomical proximity of these fungal isolates. PADs were purified from E. dentata and A. nidulans with apparent molecular masses 41 and 48 kDa, respectively. The peptide fingerprints of PADs from E. dentata and A. nidulans have been analyzed by MALDI-TOF/MS, displaying a higher sequence similarity to human PAD4 by 18% and 31%, respectively. The conserved peptide sequences of E. dentata and A. nidulans PADs displayed a higher similarity to human PAD than A. fumigatus PADs clade. PADs from both fungal isolates have an optimum pH and pH stability at 7.0-8.0, with putative pI 5.0-5.5, higher structural denaturation at pH 4.0-5.5 and 9.5-12 as revealed from absorbance at λ280nm. E. dentata PAD had a higher conformationally thermal stability than A. nidulans PAD as revealed from its lower Kr value. From the proteolytic mapping, the orientation of trypsinolytic recognition sites on the PADs surface from both fungal isolates was very similar. PADs from both isolates are calcium dependent, with participation of serine and cysteine residues on their catalytic sites. PADs displayed a higher affinity to deiminate the peptidylarginine residues with a feeble affinity to work as ADI. So, PADs from E. dentata and A. nidulans had a relatively similar conformational and kinetic properties. Further molecular modeling analysis are ongoing to explore the role of PADs in citrullination of human proteins in Aspergillosis, that will open a new avenue for unraveling the vague of protein-protein interaction of human A. nidulans pathogen.

Impeding Bacillus spore germination in vitro and in milk by soy glycinin during long cold storage.

This paper presents a study of the magnitude and mechanism of impeding the emergent spore germination of Bacillus subtilis (non-hemolytic strain) and Bacillus licheniformis (hemolytic strain) under the action of soybean glycinin and its basic subunit (BS). Incubating B. licheniformis spores with 100 µg/ml of glycinin or BS at 35°C for 24 h totally prevented the hemolytic activity associated with the emergent vegetative cells on blood agar during 3-5 days of incubation at 35°C in contrast to the control. Glycinin and BS (100 µg/ml) also resulted in the leakage of 70 and 73% of the initially germinating spore contents of A260 absorbing materials of the two bacterial species after 2 h of incubation at 35°C, respectively. Increasing the concentration of glycinin and BS up to 400 µg/ml increased the germinating spore leakage to 83 and 88%, respectively. Spore germination in Muller Hinton Broth containing glycinin and BS (100 µg/ml) was practically nil after 4 days of incubation at 35°C. Supplementing milk preheated at 80°C/5 min with both glycinin and BS kept the final vegetative cell counts down to about 30 and 20% of the corresponding control, respectively, after 50 days of storage at 4°C. Both light, and transmission electron, microscopy images revealed significant morphological and structural distorting changes in BS-treated spores of B. licheniformis.

Co-immobilization of PEGylated Aspergillus flavipes L-methioninase with glutamate dehydrogenase: a novel catalytically stable anticancer consortium.

Aspergillus flavipes L-methioninase (AfMETase) exhibits reliable pharmacokinetic properties and anticancer potency in vitro[10]. To maximize its therapeutic efficiency as protection against in vivo proteolysis, reduction of antigenicity and hyperammoniemia, the enzyme was PEGylated and coupled with glutamate dehydrogenase (GDH). The highest degree of PEGylation was measured at 40-50/1 molar ratio of PEG to AfMETase, with a lower mobility on SDS-PGE, compared to the native AfMETase. The activity of free AfMETase was reduced to 66.2% and further to 50% upon PEGylation and GDH conjugation, respectively. The highest degree of surface NH2 modification of AfMETase-GDH co-immobilizates (65%), was reported using 300 mM glutaraldehyde, with 31% methionine conversion. Using L-cysteine and L-methionine as active site protectors, the activity of PEG-AfMETase and PEG-AfMETase-GDH was increased by 14.4 and 32.9-fold, respectively. At 45°C, PEG-AfMETase, PEG-AfMETase-GDH and AfMETase-GDH conjugate have a T1/2 10.3, 8.5 and 7.6 h, inactivation rate (Kr) 0.021, 0.03 and 0.016 min, with 2.0, 1.65 and 1.47-fold stabilization, respectively. Kinetically, the three immobilizates have a relatively similar Km values for L-methionine (7.4-7.9 mM), with lower affinity to homocysteine and cysteine, with stability to PLP-enzyme inhibitors (propargylglycine and hydroxylamine), indicating the protective effect by PEG moieties on the enzyme structure. Also, the three immobilizates exhibited improved stability against proteolysis in vitro, comparing to free AfMETase.

Preservative action of 11S (glycinin) and 7S (ß-conglycinin) soy globulin on bovine raw milk stored either at 4 or 25 °C.

Considerable inhibitory antibacterial actions were exerted by the soybean 11S subunit comparable with nisin on the proliferation of total viable count, Pseudomonas count and Enterobacteriaceae count in bovine milk stored at 4 or 25 °C for 30 d and 48 h, while 7S and lysozyme were much less effective. The maximum magnitudes of bacterial reduction by 11S and nisin were in the range 2-4 log CFU/ml. The proliferation of 3 pathogenic bacteria (Listeria monocytogenes, Salmonella Enteritidis and Escherichia coli O157:H7) artificially inoculated into raw milk stored at 4 or 25 °C were particularly and significantly (P<0.05) reduced by 11S subunit and nisin (0.5% w/v), but only slightly by 7S and moderately by lysozyme. Lactose consumption, acidity development and casein degradation during storage of bovine raw milk were attenuated during storage at 4 or 25 °C and sensorial traits were better maintained by supplementation with 11S (0.5% w/v). 11S subunit may be used a safely food preservative, if permitted.

In vitro and in situ antimicrobial action and mechanism of glycinin and its basic subunit.

Glycinin, basic subunit and ß-conglycinin were isolated from soybean protein isolate and tested for their antimicrobial action against pathogenic and spoilage bacteria as compared to penicillin. The three fractions exhibited antibacterial activities equivalent to or higher than penicillin in the next order; basic subunit>glycinin>ß-conglycinin with MIC of 50, 100 and 1000 µg/mL, respectively. The IC(50%) values of the basic subunit, glycinin and ß-conglycinin against Listeria\monocytogenes were 15, 16 and 695 µg/mL, against Bacillussubtilis were 17, 20, and 612 µg/mL, and against S. Enteritidis were 18, 21 and 526 µg/mL, respectively. Transmission electron microscopy images of L. monocytogenes and S. Enteritidis exhibited bigger sizes and separation of cell wall from cell membrane when treated with glycinin or basic subunit. Scanning electron microscopy of B. subtilis indicated signs of irregular wrinkled outer surface, fragmentation, adhesion and aggregation of damaged cells or cellular debris when treated with glycinin or the basic subunits but not with penicillin. All tested substances particularly the basic subunit showed increased concentration-dependent cell permeation assessed by crystal violet uptake. The antimicrobial action of glycinin and basic subunit was swifter than that of penicillin. The cell killing efficiency was in the following descending order; basic subunit>glycinin>penicillin>ß-conglycinin and the susceptibility of the bacteria to the antimicrobial agents was in the next order: L. monocytogenes>B. Subtilis>S. Enteritidis. Adding glycinin and the basic subunit to pasteurized milk inoculated with the three bacteria; L. monocytogenes, B. Subtilis and S. Enteritidis (ca. 5 log CFU/mL) could inhibit their propagation after 16-20 days storage at 4 °C by 2.42-2.98, 4.25-4.77 and 2.57-3.01 log and by 3.22-3.78, 5.65-6.27 and 3.35-3.72 log CFU/mL, respectively.

Protective effect of whey proteins against nonalcoholic fatty liver in rats.

BACKGROUND: Nonalcoholic fatty liver disease (NAFLD) is the hepatic manifestation of the metabolic syndrome and can vary from hepatic steatosis to end-stage liver disease. It is the most common liver disease and its prevalence is increasing worldwide. In the present study, the effect of whey proteins on some parameters of NAFLD was investigated. RESULTS: Oral administration of the studied whey proteins products reduced the final body weight of rats. There was a significant reduction effect (P<0.05) of the tested proteins on hepatic triglycerides, liver enzymes (ALT and AST), lipid peroxidation (malondialdehyde level) and serum glucose. Feeding on whey proteins caused an increase in the reduced glutathione. Hepatic content of reduced glutathione was not affected by any of the used whey proteins, but it showed an increasing tendency (P>0.05). Liver histology showed an improvement of fatty infiltration in hepatocytes from whey protein groups and gives the histology of liver a normal appearance. CONCLUSIONS: The obtained results indicate a possible role for oral administration of whey proteins in the regulation of liver biochemistries in a rat's model of NAFLD. This regulatory effect of whey proteins was accompanied by an improvement in fatty infiltration in hepatocytes and a reduction of oxidative stress parameters.

Enhancing Milk Preservation with Esterified Legume Proteins.

Three methylated legume proteins; soybean protein, broad bean protein and chickpea protein as well as their respective native proteins were applied at two different concentrations (0.1 and 1%) to either raw or pasteurized milk before preservation at 4 °C for 7-14 days. Supplementation of raw milk with esterified legume proteins could ameliorate its preservation quality at 4 °C for 5 days, based on the total bacterial count (TBC) or the titratable acidity levels. Supplementing pasteurized milk with esterified legume proteins (0.1%) has significantly improved its keeping quality as it significantly reduced the total bacterial count by 3.33 and 1.80 log when preserved at 4 °C for 7 and 14 days, respectively. Esterified legume proteins (0.1%) could maintain the level of bacterial load of the pasteurized milk at its initial level of pasteurization (zero time) after 14 days of preservation at 4 °C under closed conditions.

Effectiveness of esterified whey proteins fractions against Egyptian Lethal Avian Influenza A (H5N1).

BACKGROUND: Avian influenza A (H5N1) virus is one of the most important public health concerns worldwide. The antiviral activity of native and esterified whey proteins fractions (α- lactalbumin, ß- lactoglobulin, and lactoferrin) was evaluated against A/chicken/Egypt/086Q-NLQP/2008 HPAI (H5N1) strain of clade 2.2.1 (for multiplicity of infection (1 MOI) after 72 h of incubation at 37 °C in the presence of 5% CO2) using MDCK cell lines. RESULT: Both the native and esterified lactoferrin seem to be the most active antiviral protein among the tested samples, followed by ß- lactoglobulin. α-Lactalbumin had less antiviral activity even after esterification. CONCLUSION: Esterification of whey proteins fractions especially lactoferrin and ß-lactoglobulin enhanced their antiviral activity against H5N1 in a concentration dependent manner.

Inhibition of tomato yellow leaf curl virus (TYLCV) using whey proteins.

The antiviral activity of native and esterified whey proteins fractions (alpha-lactalbumin, beta-lactoglobulin, and lactoferrin) was studied to inhibit tomato yellow leaf curl virus (TYLCV) on infected tomato plants. Whey proteins fractions and their esterified derivatives were sprayed into TYLCV-infected plants. Samples were collected from infected leaves before treatment, 7 and 15 days after treatment for DNA and molecular hybridization analysis. The most evident inhibition of virus replication was observed after 7 and 15 days using alpha-lactoferrin and alpha-lactalbumin, respectively. Native and esterified lactoferrin showed complete inhibition after 7 days. On the other hand, native beta-lactoglobulin showed inhibition after 7 and 15 days whereas esterified beta-lactoglobulin was comparatively more effective after 7 days. The relative amount of viral DNA was less affected by the esterified alpha-lactalbumin whereas native alpha-lactalbumin inhibited virus replication completely after 15 days. These results indicate that native or modified whey proteins fractions can be used for controlling the TYLCV-infected plants.