Search results for: whey protein

Authors: Phakamas Rachamontree, Theerawut Phusantisampan, Natthakorn Woravutthikul, Peerapong Pornwongthong, Malinee Sriariyanun

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A total of 115 yeast strains isolated from local cassava processing wastes were measured for crude protein content. Among these strains, the strain MSY-2 possessed the highest protein concentration (>3.5 mg protein/mL). By using molecular identification tools, it was identified to be a strain of Pichia kudriavzevii based on similarity of D1/D2 domain of 26S rDNA region. In this study, to optimize the protein production by MSY-2 strain, Response Surface Methodology (RSM) was applied. The tested parameters were the carbon content, nitrogen content, and incubation time. Here, the value of regression coefficient (R2) = 0.7194 could be explained by the model, which is high to support the significance of the model. Under the optimal condition, the protein content was produced up to 3.77 g per L of the culture and MSY-2 strain contain 66.8 g protein per 100 g of cell dry weight. These results revealed the plausibility of applying the novel strain of yeast in single-cell protein production.

Keywords: single cell protein, response surface methodology, yeast, cassava processing waste

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Authors: Seyed Mohammad Hosein Al Omrani Nejad, Ali Rezvani Aghdam

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This study was done in order to evaluation different irrigation intervals on amount of protein, and gel production in Aloe vera, a traditional medicinal plant. Plants was plnted in Greenhouse and irrigated according to Accumulative Pan Evaporation(APE). The treatments were included 20, 40, 60, 80, 100, 120, 140, 160, 180, and 200 mm APE which has been showed W1,W2, W3, W4, W5, W6, W7, W8,W9 and W10 respectively.The amount of protein and gel production was measured seperately. Results showed that highest protein and fresh weight of gel obtained plants which irrigated W6 and W7 respectively. According to these results can recomend which if plant irrigatedwhen APE reached 120 and 140 mm by Class A Evaporation Pan method gel production and protein would besuitable in north of khozestan province in limited irrigation conditions.

Keywords: irrigation, protein, gel, aloe vera, Iran

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Authors: Er-Yuan Chuang

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Bio-mimetic matters have biological functionalities. This might be valuable in the development of versatile biomaterials. At biological fields, protein-based materials might be components to form a 3D network of extracellular biomolecules, containing growth factors. Also, the protein-based biomaterial provides biochemical and structural assistance of adjacent cells. In this study, we try to prepare protein based biomaterial, which was harvested from living animal. We analyzed it’s chemical, physical and biological property in vitro. Besides, in vivo bio-interaction of the prepared biomimetic matrix was tested in an animal model. The protein-based biomaterial has degradability and biocompatibility. This development could be used for tissue regenerations and be served as platform technologies.

Keywords: protein based, in vitro study, in vivo study, biomaterials

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Authors: Kersch-Counet C., Fransen K. H. S., Broyd M., Nyakayiru J. D. O. A., Schoemaker M. H., Mallee L. F., Bovee-Oudenhoven I. M. J.

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Background: Around one third of people, including athletes, suffer from feelings of gut discomfort. Fermentation of dairy is a process that has been associated with products that can improve gut health. However, insight in (potential) health benefits of most fermented foods is limited to chemical analyses and in-vitro models. Objective: The aim of this open-label, single-arm explorative trial was to investigate in a real life setting the effect of consumption of a fermented whey product for 3 weeks on self-perceived physical and mental wellbeing and digestive issues in 150 US recreational athletes (20-50 years of age) with self-reported gut complaints at enrolment. Methods: Participants living at the West-Coast of the US received for 3 weeks a daily powder of 15 g of BiotisTM Fermentis to be mixed in water using a supplied shaker. Weekly questionnaires were conducted by MMR research to study the effect on physical/mental health issues and self-perceived gut complaints. Non-parametric tests (e.g., Friedman test) were used to assess statistical differences over time while the Kruskal-Wallis and Wilcoxon signed-rank tests were used for sub-groups analysis. Results: Bloating, stress and anxiety were the top 3 issues of the US recreational athletes. Satisfaction of physical wellbeing increased significantly throughout the 3-weeks of fermented whey product consumption (p<0.0005). Combined digestive issues decreased significantly after 2- and 3-weeks of product consumption, with bloating showing a significant reduction (p<0.05). There was a trend that self-reported stress levels reduced after 3 weeks and participants said to significantly feel more active, energetic, and vital (p<0.05). Subgroup analysis showed that gender and habitual protein supplement consumption were associated with specific health issues and modulated the response to the fermented dairy product. Conclusion: Daily consumption of the fermented BiotisTM Fermentis product is associated with a reduction in self-perceived gastrointestinal symptoms and improved overall wellbeing and mood state in US recreational athletes. This large nutrition and health consumer study brings valuable insights in self-reported gut complaints of recreational athletes in the US and their response to a fermented dairy product. A controlled clinical trial in a targeted population is recommended to scientifically substantiate the product effect as observed in this explorative study.

Keywords: real-life study, digestive health, fermented whey, sports

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Authors: Mohamed Abdullah Ahmed

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In a study of chickpea protein isolate (CPI) preparation, the wet alkaline extraction was carried out. The objectives were to determine the optimal extracting conditions of CPI and apply CPI into a sponge cake recipe to replace egg and make acceptable product. The design used in extraction was a central composite design. The response surface methodology was preferred to graphically express the relationship between extraction time and pH with the output variables of percent yield and protein content of CPI. It was noted that optimal extracting conditions were 60 min and pH 10.5 resulting in 90.07% protein content and 89.15% yield of CPI. The protein isolate (CPI) could be incorporated in cake to 20% without adversely affecting the cake physical properties such as cake hardness and sensory attributes. The higher protein content in cake was corresponding to the amount of CPI added. Therefore, adding CPI can significantly (p<0.05) increase protein content in cake. However, sensory evaluation showed that adding more than 20% of CPI decreased the overall acceptability. The results of this investigation could be used as a basic knowledge of CPI utilization in other food products.

Keywords: chick bean protein isolate, sponge cake, utilization, sponge

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Authors: Shayli Varasteh Moradi, Wayne A. Johnston, Dejan Gagoski, Kirill Alexandrov

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The demand for a rapid and more efficient technique to identify protein-protein interaction particularly in the areas of therapeutics and diagnostics development is growing. The method described here is a rapid in vitro protein-protein interaction analysis approach based on AlphaLISA technology combined with Leishmania tarentolae cell-free protein production (LTE) system. Cell-free protein synthesis allows the rapid production of recombinant proteins in a multiplexed format. Among available in vitro expression systems, LTE offers several advantages over other eukaryotic cell-free systems. It is based on a fast growing fermentable organism that is inexpensive in cultivation and lysate production. High integrity of proteins produced in this system and the ability to co-express multiple proteins makes it a desirable method for screening protein interactions. Following the translation of protein pairs in LTE system, the physical interaction between proteins of interests is analysed by AlphaLISA assay. The assay is performed using unpurified in vitro translation reaction and therefore can be readily multiplexed. This approach can be used in various research applications such as epitope mapping, antigen-antibody analysis and protein interaction network mapping. The intra-viral protein interaction network of Zika virus was studied using the developed technique. The viral proteins were co-expressed pair-wise in LTE and all possible interactions among viral proteins were tested using AlphaLISA. The assay resulted to the identification of 54 intra-viral protein-protein interactions from which 19 binary interactions were found to be novel. The presented technique provides a powerful tool for rapid analysis of protein-protein interaction with high sensitivity and throughput.

Keywords: AlphaLISA technology, cell-free protein expression, epitope mapping, Leishmania tarentolae, protein-protein interaction

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Authors: Duc Dong Do, Ngoc Ha Tran, Thanh Hai Dang, Cao Cuong Dang, Xuan Huan Hoang

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Global aligning two protein-protein interaction networks is an essentially important task in bioinformatics/computational biology field of study. It is a challenging and widely studied research topic in recent years. Accurately aligned networks allow us to identify functional modules of proteins and/ororthologous proteins from which unknown functions of a protein can be inferred. We here introduce a novel efficient heuristic global network alignment algorithm called FASTAn, including two phases: the first to construct an initial alignment and the second to improve such alignment by exerting a local optimization repeated procedure. The experimental results demonstrated that FASTAn outperformed the state-of-the-art global network alignment algorithm namely SPINAL in terms of both commonly used objective scores and the run-time.

Keywords: FASTAn, Heuristic algorithm, biological network alignment, protein-protein interaction networks

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Authors: Xiao Zhou, Jianlin Cheng

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A single amino acid mutation can have a significant impact on the stability of protein structure. Thus, the prediction of protein stability change induced by single site mutations is critical and useful for studying protein function and structure. Here, we presented a deep learning network with the dropout technique for predicting protein stability changes upon single amino acid substitution. While using only protein sequence as input, the overall prediction accuracy of the method on a standard benchmark is >85%, which is higher than existing sequence-based methods and is comparable to the methods that use not only protein sequence but also tertiary structure, pH value and temperature. The results demonstrate that deep learning is a promising technique for protein stability prediction. The good performance of this sequence-based method makes it a valuable tool for predicting the impact of mutations on most proteins whose experimental structures are not available. Both the downloadable software package and the user-friendly web server (DNpro) that implement the method for predicting protein stability changes induced by amino acid mutations are freely available for the community to use.

Keywords: bioinformatics, deep learning, protein stability prediction, biological data mining

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Authors: Duygu Çimen, Nilay Bereli, Adil Denizli

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In this study is to synthesis magnetic nanoparticles for purify protein C. For this aim, N-Methacryloyl-(L)-histidine methyl ester (MAH) containing 2-hydroxyethyl methacrylate (HEMA) based magnetic nanoparticles were synthesized by using micro-emulsion polymerization technique for templating protein C via metal chelation. The obtained nanoparticles were characterized with Fourier transform infrared spectroscopy (FTIR), transmission electron microscopy (TEM), zeta-size analysis and electron spin resonance (ESR) spectroscopy. After that, they were used for protein C purification from aqueous solution to evaluate/optimize the adsorption condition. Hereby, the effecting factors such as concentration, pH, ionic strength, temperature, and reusability were evaluated. As the last step, protein C was determined with sodium dodecyl sulfate-polyacrylamide gel electrophoresis.

Keywords: immobilized metal affinity chromatography (IMAC), magnetic nanoparticle, protein C, hydroxyethyl methacrylate (HEMA)

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Authors: Ahmed M. Hjazi, Bader M. Hjazi

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Protein 4.1 is a crucial component of complex interactions between the cytoskeleton and other junctional complex proteins. When the gene encoding this protein is altered, resulting in reduced expression, or when the protein is absent, the red cell undergoes a significant structural change. This research aims to achieve a deeper comprehension of the biochemical effects of red cell protein deficiency. A Tandem Mass Spectrometry Analysis (TMT-MS/MS) of patient cells lacking protein 4.1 compared to three healthy controls was achieved by the Proteomics Institute of the University of Bristol. The SDS-PAGE and Western blotting were utilized on the original patient sample and controls to partially confirm TMT MS/MS data analysis of the protein-4.1-deficient cells. Compared to healthy controls, protein levels in samples lacking protein 4.1 had a significantly higher concentration of proteins that probably originated from reticulocytes. This could occur if the patient has an elevated reticulocyte count. The increase in chaperone and reticulocyte-associated proteins was most notable in this study. This may result from elevated quantities of reticulocytes in patients with hereditary elliptocytosis.

Keywords: hereditary elliptocytosis, protein 4.1, red cells, tandem mass spectrometry data.

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Authors: Yusuf Ali Abdulle, Azhar Uddin Keerio

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Background: Protein elicitors play a key role in signaling or displaying plant defense mechanisms and emerging as vital tools for bio-control of insects. This study was aimed at the characterization of the novel protein elicitor isolated from entomopathogenic fungi Lecanicillium lecanii (V3) strain and its activity against Whitefly, Bemisia tabaci in cotton. The sequence of purified elicitor protein showed 100% similarity with hypothetical protein LEL_00878 [Cordyceps confragosa RCEF 1005], GenBank no (OAA81333.1). This novel protein elicitor has 253 amino acid residues and 762bp with a molecular mass of 29 kDa. The protein recombinant was expressed in Escherichia coli using pET‐28a (+) plasmid. Effects of purified novel protein elicitor on Bemisia tabaci were determined at three concentrations of protein (i.e., 58.32, 41.22, 35.41 μg mL⁻¹) on cotton plants and were exposed to newly molted adult B.tabaci. Bioassay results showed a significant effect of the exogenous application of novel protein elicitor on B. tabaci in cotton. In addition, the gene expression analysis found a significant up-regulation of the major genes associated with salicylic acid (SA) and jasmonic acid (JA) linked plant defense pathways in elicitor protein-treated plants. Our results suggested the potential application of a novel protein elicitor derived from Lecanicillium lecanii as a future bio-intensive controlling approach against the whitefly, Bemisia tabaci.

Keywords: resistance, Lecanicillium lecanii, secondary metabolites, whitefly

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Authors: Amr A. El Hanafy, Muhammad I. Qureshi, Jamal Sabir, Mohamed Mutawakil, Mohamed M. Ahmed, Hassan El Ashmaoui, Hassan Ramadan, Mohamed Abou-Alsoud, Mahmoud Abdel Sadek

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β-Lactoglobulin (β-LG) is the dominant non-casein whey protein found in bovine milk and of most ruminants. The amino acid sequence of β-LG along with its 3-dimensional structure illustrates linkage with the lipocalin superfamily. Preliminary studies in goats indicated that milk yield can be influenced by polymorphism in genes coding for whey proteins. The aim of this study is to identify and evaluate the incidence of functional polymorphisms in the exonic and intronic portions of β-LG gene in native Saudi goat breeds (Ardi, Habsi, and Harri). Blood samples were collected from 300 animals (100 for each breed) and genomic DNA was extracted using QIAamp DNA extraction Kit. A fragment of the β-LG gene from exon 7 to 3’ flanking region was amplified with pairs of specific primers. Subsequent digestion with Sac II restriction endonuclease revealed two alleles (A and B) and three different banding patterns or genotypes i.e. AA, AB and BB. The statistical analysis showed that β-LG AA genotype had higher milk yield than β-LG AB and β-LG BB genotypes. Nucleotide sequencing of the selected β-LG fragments was done and submitted to GenBank NCBI (Accession No. KJ544248, KJ588275, KJ588276, KJ783455, KJ783456 and KJ874959). Two already established SNPs in exon 7 (+4601 and +4603) and one fresh SNP in the 3’ UTR region were detected in the β-LG fragments with designated AA genotype. The polymorphisms in exon 7 did not produce any amino acid change. Phylogenetic analysis on the basis of nucleotide sequences of native Saudi goats indicated evolutional similarity with the GenBank reference sequences of goat, Bubalus bubalis and Bos taurus.

Keywords: β-Lactoglobulin, Saudi goats, PCR-RFLP, functional polymorphism, nucleotide sequencing, phylogenetic analysis

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Authors: Angela U. Makolo, Temitayo A. Olagunju

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The knowledge of signaling pathways is central to understanding the biological mechanisms of organisms since it has been identified that in eukaryotic organisms, the number of signaling pathways determines the number of ways the organism will react to external stimuli. Signaling pathways are studied using protein interaction networks constructed from protein-protein interaction data obtained using high throughput experimental procedures. However, these high throughput methods are known to produce very high rates of false positive and negative interactions. In order to construct a useful protein interaction network from this noisy data, computational methods are applied to validate the protein-protein interactions. In this study, a computational technique to identify signaling pathways from a protein interaction network constructed using validated protein-protein interaction data was designed. A weighted interaction graph of the Saccharomyces cerevisiae (Baker’s Yeast) organism using the proteins as the nodes and interactions between them as edges was constructed. The weights were obtained using Bayesian probabilistic network to estimate the posterior probability of interaction between two proteins given the gene expression measurement as biological evidence. Only interactions above a threshold were accepted for the network model. A pathway was formalized as a simple path in the interaction network from a starting protein and an ending protein of interest. We were able to identify some pathway segments, one of which is a segment of the pathway that signals the start of the process of meiosis in S. cerevisiae.

Keywords: Bayesian networks, protein interaction networks, Saccharomyces cerevisiae, signalling pathways

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Authors: Andrea Vasquez, Pedro Maldonado-Alvarado

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In general, 'gluten-free' foods like breadmaking products provide functional or nutraceutical benefits for the consumer's health and increased their demand on the market. In Ecuador, there is an overproduction of superfoods, and the food by-products are undervalued. For the first time, to the author's best knowledge, gluten-free bread mixtures from quinoa and banana flour, cassava starch, lupine flour (LF), or whey protein (WP) with hydroxypropylmethylcellulose (HPMC) and transglutaminase (TG) were evaluated on their functional and sensory properties. Free amino groups and thiols, rheology, and electrophoresis SDS PAGE were performed to analyze the crosslinking of TG at different concentrations with HC or PL proteins. Dough characterization, pasting properties were evaluated, respectively, by a MIXOLAB and a rheometer with a pasting cell. The texture, porosity, and loaf volume were characterized using a texturometer, ImageJ software, and breadmaking ability, respectively. Finally, a breadmaking aptitude and sensorial bread acceptability were performed. A significant decrease in the content of free amino groups (0.16 to 0.11 and 0.46 to 0.36 mM/mg of protein) and free thiol groups (0.37 to 0.21 and 1.79 to 1.32 mM/mg protein) was observed when 1.0% and 0.5% TG were added to LF and WP, respectively. In apparent viscosity analysis, the action of TG on HC proteins changes their viscosity, while the viscosity of LF is not modified by TG. Results of electrophoresis in PL showed bands of higher molecular weight of different fragments of proteins with 1% TG. Formulation with 59.8, 39.9, 160.8, 6.0, 1.0, and 1.5% of, respectively, QF, BF, CS, LF or WP, TG, and HPMC had the best properties in dough parameters, pasting parameters (lower pasting temperature and higher peak viscosity), best crumb structure, lower crumb hardness and higher loaf volume (2.24 and 2.28 mL/g). All the loaves of bread were acceptable in baking aptitude and general acceptability.

Keywords: breadmaking, gluten-free, superfoods, by-products, Ecuador

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Authors: Gül Ebru Orhun

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A half diallel crossing design was carried out during 2011 and 2012 growing seasons under Çanakkale-Turkey ecological conditions. In this research, 20 F1 maize hybrids obtained by 6x6 half diallel crossing were used. Gene action for protein content and grain yield traits were explored in half set involving six elite inbred lines. According to the results diallel analysis dominance and additive gene variances were determined for protein content. Variance/Co-variance graphs revealed for grain yield and protein content traits. In this study, inheritance of grain yield and protein content demonstrated over-dominance type of gene action.

Keywords: protein, maize, inheritance, gene action

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Authors: Ayobami Adeyemo, Michael Chimonyo, Munyaradzi Marufu

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Gastrointestinal nematode (GNT) infection significantly hinder sustainable and profitable sheep production on rangelands. While vitamin E and protein supplementation have individually proven to improve host immunity to parasitism in lambs, to our knowledge, there is no information on the interaction of dietary vitamin E and protein supplementation on lamb growth and GIN faecal egg counts in naturally infected lambs. Therefore, the current study investigated the interaction of dietary protein and vitamin E supplementation on faecal egg counts (FEC) and growth performance of lambs. Twenty four Dohne Merino lambs aged 12 months were allocated equally to each of four treatment combinations, with six lambs in each treatment group for a period of eight weeks. Treatment one lambs received dietary protein and vitamin E (PE), treatment two lambs received dietary protein and no vitamin E (PNE), treatment three received dietary vitamin E and no protein (NPE), and treatment four received no dietary protein and vitamin E supplementation (NPNE). The lambs were allowed to graze on Pennisetum clandestinum contaminated with a heavy load of nematodes. Dietary protein supplementation increased (P < 0.01) average daily gain (ADG) and body condition scores (BCS). Dietary vitamin E supplementation had no effect (P > 0.05) on ADG and BCS. There was no interaction (P > 0.05) between dietary protein and vitamin E supplementation on ADG and BCS. Combined supplementation of dietary protein and vitamin E supplementation significantly reduced (P < 0.01) faecal egg counts and larval counts, respectively. Also, dietary protein and vitamin E supplementation reduced GNT faecal egg counts over the exposure period. The current findings support the hypothesis that the interaction of dietary protein and vitamin E supplementation reduced faecal egg counts and larval counts in lambs. This necessitates future findings on the interaction of dietary protein and vitamin E supplementation on blood associated profiles.

Keywords: gastrointestinal nematodes, nematode eggs, Haemonchus, Trichostrongylus

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Authors: Jin Choi, Zahra Aminikhoei, Yi-Oh Kim, Sang-Min Lee

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A 4 × 2 factorial experiment was conducted to determine the optimum dietary protein and lipid levels for juvenile fancy carp, Cyprinus carpio var. koi. Eight experimental diets were formulated to contain four protein levels (200, 300, 400, and 500 g kg-1) with two lipid levels (70 and 140 g kg-1). Triplicate groups of fish (initial weight, 12.1±0.2 g fish-1) were hand-fed the diets to apparent satiation for 8 weeks. Weight gain, daily feed intake, feed efficiency ratio and protein efficiency ratio were significantly (P < 0.0001) affected by dietary protein level, but not by dietary lipid level (P > 0.05). Weight gain and feed efficiency ratio tended to increase as dietary protein level increased up to 400 and 500 g kg-1, respectively. Daily feed intake of fish decreased with increasing dietary protein level and that of fish fed diet contained 500 g kg-1 protein was significantly lower than other fish groups. The protein efficiency ratio of fish fed 400 and 500 g kg-1 protein was lower than that of fish fed 200 and 300 g kg-1 protein. Moisture, crude protein and crude lipid contents of muscle and liver were significantly affected by dietary protein, but not by dietary lipid level (P > 0.05). The increase in dietary lipid level resulted in an increase in linoleic acid in liver and muscle paralleled with a decrease in n-3 highly unsaturated fatty acids content in muscle of fish. In considering these results, it was concluded that the diet containing 400 g kg-1 protein with 70 g kg-1 lipid level is optimal for growth and efficient feed utilization of juvenile fancy carp.

Keywords: fancy carp, dietary protein, dietary lipid, Cyprinus carpio, fatty acid

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Authors: Muhammad H. Alu’datt, Inteaz Alli

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This study was conducted to prepare and evaluate the biological properties of protein co-precipitates from flaxseed and soybean. Protein was prepared by NaOH extraction through the mixing of soybean flour (Sf) and flaxseed flour (Ff) or mixtures of soybean extract (Se) and flaxseed extract (Fe). The protein co-precipitates were precipitated by isoelectric (IEP) and isoelectric-heating (IEPH) co-precipitation techniques. Effects of extraction and co-precipitation techniques on co-precipitate yield were investigated. Native-PAGE, SDS-PAGE were used to study the molecular characterization. Content and antioxidant activity of extracted free and bound phenolic compounds were evaluated for protein co-precipitates. Removal of free and bound phenolic compounds from protein co-precipitates showed little effects on the electrophoretic behavior of the proteins or the protein subunits of protein co-precipitates. Results showed that he highest protein contents and yield were obtained in for Sf-Ff/IEP co-precipitate with values of 53.28 and 25.58% respectively as compared to protein isolates and other co-precipitates. Results revealed that the Sf-Ff/IEP showed a higher content of bound phenolic compounds (53.49% from total phenolic content) as compared to free phenolic compounds (46.51% from total phenolic content). Antioxidant activities of extracted bound phenolic compounds with and without heat treatment from Sf-Ff/IEHP were higher as compared to free phenolic compounds extracted from other protein co-precipitates (29.68 and 22.84%, respectively).

Keywords: antioxidant, phenol, protein co-precipitate, yield

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Authors: Jannika Dombrowski, Sabine Ambros, Ulrich Kulozik

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Due to the increasing popularity of healthy products, probiotics are still of rising importance in food manufacturing. With the aim to amplify the field of probiotic application to non-chilled products, the cultures have to be preserved by drying. Microwave drying has proved to be a suitable technique to achieve relatively high survival rates, resulting from drying at gentle temperatures, among others. However, diffusion limitation due to compaction of cell suspension during drying can prolong drying times as well as deteriorate product properties (grindability, rehydration performance). Therefore, we aimed to embed probiotics in an aerated matrix of whey proteins (surfactants) and di-/polysaccharides (foam stabilization, probiotic protection) during drying. As a result of the manifold increased inner surface of the cell suspension, drying performance was enhanced significantly as compared to non-foamed suspensions. This work comprises investigations on suitable foam matrices, being stable under vacuum (variation of protein concentration, type and concentration of di-/polysaccharide) as well as development of an applicable microwave drying process in terms of microwave power, chamber pressure and maximum product temperatures. Performed analyses included foam characteristics (overrun, drainage, firmness, bubble sizes), and properties of the dried cultures (survival, activity). In addition, efficiency of the drying process was evaluated.

Keywords: foam structure, microwave drying, polysaccharides, probiotics

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Authors: Saleh H. Salmen, Sulaiman Ali Alharbi, Hany M. Yehia, Mohammad A. Khiyami, Milton Wainwright, Naiyf S. Alharbi, Arunachalam Chinnathambi

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An antifungal protein synthesized by Bacillus cereus has been partially purified by the use of ammonium sulfate precipitation and Sephadex-G-200 column chromatography. The protein was produced from Bacillus cereus grown in potato Dextrose Broth Medium (PDB) at 30 ºC for 3 days at 100 rpm. The protein showed antagonistic effect against some fungi and yeasts. Crude extract from medium and semi-purified protein were tested in vitro against both fungi and yeasts using the disc diffusion method in order to detect the inhibitory effect of the protein. Zones of inhibition of the following diameter were found (mm) were Alternaria alternate (28), Rhodotorula glutinis (20), Fusarium sp. (16), Rhizopus sp. (15), Penicillium digitatum (13), Mucor sp. (13) and Aspergillus niger (10). The isolated protein was found to have a molecular weight of ~35kDa by sodium deodecyl sulfate-poly acrylamide gel electrophoresis. The data showed that the protein of Bacillus cereus has antifungal activity, a fact which points to the possibility of using it as a bio-control agent against some fungi, findings which emphasize the potential role of B. cereus as an important bio-control agent.

Keywords: bacillus cereus, ~35kDa protein, molds, yeasts

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Authors: Carlos A. C. G. Neto, Natan C. G. e Silva , Thaís de O. Costa, Luciana R. B. Gonçalves, Maria V. P. Rocha

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β-galactosidases (E.C. 3.2.1.23) are enzymes that have attracted by catalyzing the hydrolysis of lactose and in producing galacto-oligosaccharides by favoring transgalactosylation reactions. These enzymes, when immobilized, can have some enzymatic characteristics substantially improved, and the coating of supports with multifunctional polymers is a promising alternative to enhance the stability of the biocatalysts, among which polyethylenimine (PEI) stands out. PEI has certain properties, such as being a flexible polymer that suits the structure of the enzyme, giving greater stability, especially for multimeric enzymes such as β-galactosidases. Besides that, protects them from environmental variations. The use of chitosan support coated with PEI could improve the catalytic efficiency of β-galactosidase from Kluyveromyces lactis in the transgalactosylation reaction for the production of prebiotics, such as lactulose since this strain is more effective in the hydrolysis reaction. In this context, the aim of the present work was first to develop biocatalysts of β-galactosidase from K. lactis immobilized on chitosan-coated with PEI, determining the immobilization parameters, its operational and thermal stability, and then to apply it in hydrolysis and transgalactolisation reactions to produce lactulose using whey as a substrate. The immobilization of β-galactosidase in chitosan previously functionalized with 0.8% (v/v) glutaraldehyde and then coated with 10% (w/v) PEI solution was evaluated using an enzymatic load of 10 mg protein per gram support. Subsequently, the hydrolysis and transgalactosylation reactions were conducted at 50 °C, 120 RPM for 20 minutes, using whey supplemented with fructose at a ratio of 1:2 lactose/fructose, totaling 200 g/L. Operational stability studies were performed in the same conditions for 10 cycles. Thermal stabilities of biocatalysts were conducted at 50 ºC in 50 mM phosphate buffer, pH 6.6 with 0.1 mM MnCl2. The biocatalyst whose support was coated was named CHI_GLU_PEI_GAL, and the one that was not coated was named CHI_GLU_GAL. The coating of the support with PEI considerably improved the parameters of immobilization. The immobilization yield increased from 56.53% to 97.45%, biocatalyst activity from 38.93 U/g to 95.26 U/g and the efficiency from 3.51% to 6.0% for uncoated and coated support, respectively. The biocatalyst CHI_GLU_PEI_GAL was better than CHI_GLU_GAL in the hydrolysis of lactose and production of lactulose, converting 97.05% of lactose at 5 min of reaction and producing 7.60 g/L lactulose in the same time interval. QUI_GLU_PEI_GAL biocatalyst was stable in the hydrolysis reactions of lactose during the 10 cycles evaluated, converting 73.45% lactose even after the tenth cycle, and in the lactulose production was stable until the fifth cycle evaluated, producing 10.95 g/L lactulose. However, the thermal stability of CHI_GLU_GAL biocatalyst was superior, with a half-life time 6 times higher, probably because the enzyme was immobilized by covalent bonding, which is stronger than adsorption (CHI_GLU_PEI_GAL). Therefore, the strategy of coating the supports with PEI has proven to be effective for the immobilization of β-galactosidase from K. lactis, considerably improving the immobilization parameters, as well as, the catalytic action of the enzyme. Besides that, this process can be economically viable due to the use of an industrial residue as a substrate.

Keywords: β-galactosidase, immobilization, kluyveromyces lactis, lactulose, polyethylenimine, transgalactosylation reaction, whey

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Authors: Nais Pinta Adetya, H. Hadiyanto

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Microalgae has a potential to be utilized as food and natural colorant. The microalgae components consists of three main parts, these are lipid, protein, and carbohydrate. Crucial step in producing lipid and protein from microalgae is extraction. Microalgae has high water level (70-90%), it causes drying process of biomass needs much more energy and also has potential to distract lipid and protein from microalgae. Extraction of lipid from wet biomass is able to take place efficiently with cell disruption of microalgae by osmotic shock method. In this study, osmotic shock method was going to be integrated with ultrasound to maximalize the extraction yield of lipid and protein from wet biomass Spirulina sp. with osmotic shock method assisted ultrasound. This study consisted of two steps, these were osmotic shock process toward wet biomass and ultrasound extraction assisted. NaCl solution was used as osmotic agent, with the variation of concentrations were 10%, 20%, and 30%. Extraction was conducted in 40°C for 20 minutes with frequency of ultrasound wave was 40kHz. The optimal yield of protein (2.7%) and (lipid 38%) were achieved at 20% osmotic agent concentration.

Keywords: extraction, lipid, osmotic shock, protein, ultrasound

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Authors: Jer-Yuh Liu, Je-Chiuan Ye, Jin-Ming Hwang

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Protein kinase C alpha (PKCα) is a key signaling molecule in human cancer development. As a therapeutic strategy, targeting PKCα is difficult because the molecule is ubiquitously expressed in non-malignant cells. PKCα is regulated by the cooperative interaction of the transcription factors myeloid zinc finger 1 (MZF-1) and Ets-like protein-1 (Elk-1) in human cancer cells. By conducting tissue array analysis, herein, we determined the protein expression of MZF-1/Elk-1/PKCα in various cancers. The data show that the expression of MZF-1/Elk-1 is correlated with that of PKCα in hepatocellular carcinoma (HCC), but not in bladder and lung cancers. In addition, the PKCα down-regulation by shRNA Elk-1 was only observed in the HCC SK-Hep-1 cells. Blocking the interaction between MZF-1 and Elk-1 through the transfection of their binding domain MZF-160–72 decreased PKCα expression. This step ultimately depressed the epithelial-mesenchymal transition potential of the HCC cells. These findings could be used to develop an alternative therapeutic strategy for patients with the PKCα-derived HCC.

Keywords: protein kinase C alpha, myeloid zinc finger 1, ets-like protein-1, hepatocellular carcinoma

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Authors: Hatairuk Tungkasen, Somrudee Phetchrid, Suwapat Jaidee, Supinya Yoomak, Chantana Kankamol, Mayuree Pumipaiboon, Mayuva Areekijseree

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Ovaries of reproductive female pigs were obtained from local slaughterhouses in Nakorn Pathom Province, Thailand. Follicular fluid of medium follicle (5-6 diameters) and large follicles (7-8 mm and 10 mm in diameter) were aspirated and collected by sterile technique and analyzed protein pattern. The follicular fluid protein bands were found by SDS-PAGE which has no protein band in difference compared to standard protein band. So we chose protein band molecular weight 50, 62-65, 75-80, 90, 120-160, and >220 kDa were analyzed by LC/MS/MS. The result was found immunoglobulin gamma chain, keratin, transferrin, heat shock protein, and plasminogen precursor, ceruloplasmin, and hemopexin, and protease, respectively. All proteins play important roles in promotion and regulation on growth and development of reproductive cells. The result of this study found many proteins which were useful and important for in vitro oocyte maturation and embryonic development of cell technology in animals. The further study will be use porcine follicular fluid protein of medium and large follicles as feeder cells in in vitro condition to promote oocyte and embryo maturation.

Keywords: follicular fluid protein, LC/MS/MS, porcine oocyte, SDS-PAGE

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Authors: Birgul Hizlar, Sibel Karakaya

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Recently, increasing attention has been given to carotenoid bioaccessibility and bioavailability in the field of nutrition research. As a consequence of their lipophilic nature and their specific localization in plant-based tissues, carotenoid bioaccessibility and bioavailability is generally quite low in raw fruits and vegetables, since carotenoids need to be released from the cellular matrix and incorporated in the lipid fraction during digestion before being absorbed. Today’s approach related to improving the bioaccessibility is to design food matrix. Recently, the newest approach, excipient food, has been introduced to improve the bioavailability of orally administered bioactive compounds. The main idea is combining food and another food (the excipient food) whose composition and/or structure is specifically designed for improving health benefits. In this study, effects of food processing, food matrix and the addition of excipient foods on the carotenoid bioaccessibility of carrots were determined. Different excipient foods (olive oil, lemon juice and whey curd) and different food matrices (grating, boiling and mashing) were used. Total carotenoid contents of the grated, boiled and mashed carrots were 57.23, 51.11 and 62.10 μg/g respectively. No significant differences among these values indicated that these treatments had no effect on the release of carotenoids from the food matrix. Contrary to, changes in the food matrix, especially mashing caused significant increase in the carotenoid bioaccessibility. Although the carotenoid bioaccessibility was 10.76% in grated carrots, this value was 18.19% in mashed carrots (p<0.05). Addition of olive oil and lemon juice as excipients into the grated carrots caused 1.23 times and 1.67 times increase in the carotenoid content and the carotenoid bioaccessibility respectively. However, addition of the excipient foods in the boiled carrot samples did not influence the release of carotenoid from the food matrix. Whereas, up to 1.9 fold increase in the carotenoid bioaccessibility was determined by the addition of the excipient foods into the boiled carrots. The bioaccessibility increased from 14.20% to 27.12% by the addition of olive oil, lemon juice and whey curd. The highest carotenoid content among mashed carrots was found in the mashed carrots incorporated with olive oil and lemon juice. This combination also caused a significant increase in the carotenoid bioaccessibility from 18.19% to 29.94% (p<0.05). When compared the results related with the effect of the treatments on the carotenoid bioaccessibility, mashed carrots containing olive oil, lemon juice and whey curd had the highest carotenoid bioaccessibility. The increase in the bioaccessibility was approximately 81% when compared to grated and mashed samples containing olive oil, lemon juice and whey curd. In conclusion, these results demonstrated that the food matrix and addition of the excipient foods had a significant effect on the carotenoid content and the carotenoid bioaccessibility.

Keywords: carrot, carotenoids, excipient foods, food matrix

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Authors: Hanan Azzam1, Nashwa Abousamra1, Amany Mansour1, Yaser Abd El-dayem2, , Solafa Elsharawy1

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Background: Inherited thrombophilias are a heterogenous group of conditions which have been implicated in a variety of pregnancy complications. More recently, deficiency of protein Z (PZ) has been liked to pregnancy complications, including preterm delivery. Aim: We designed this study to evaluate the association of inherited thrombophilias including [Protein C (PC), Protein S (PS), Anti thrombin III (ATIII) deficiency and activated protein C (APC) resistance] and protein Z deficiency with a variety of pregnancy complications. Patients and Methods: 60 women with different pregnancy complications, including 20 patients with preeclampsia, 20 patients with intrauterine growth resistance (IUGR), and 20 patients with intrauterine fetal death (IUFD), in addition to 30 healthy pregnant women were recruited for the present study. PC and free PS antigen, ATIII activity, modified functional APC-resistance, and PZ levels were determined. Results: There was no significant association between inherited thrombophilias and complicated pregnancies as regards PC deficiency (p=1.0), AT III and PS deficiency (p=0.312), and APC-resistance (P=0.083). PZ was significantly associated with complicated pregnancies (p=0.012). Patients with protein Z levels below 1.5 µg/ml were considered deficient. Accordingly, we demonstrated protein Z deficiency in 30% of complicated pregnancies (RR 6.0, 95% CI 1.29-27.90;p=0.022), 20% of preeclampsia (RR 3.5, 95% CI 0.57 – 21.28; P = 0.174), 40% of IUGR (RR 9.3 95% CI 1.72-50.61; P = 0.010) and 30% of IUFD (RR 6, 95% CI 1.07 – 33.64; P = 0.042). Conclusions: These findings indicate the absence of association of inherited thrombophilias, including PC, PS, AT III deficiency, and APC resistance with pregnancy complications. However, PZ deficiency is associated with increased risk of pregnancy complications, especially intrauterine growth restriction and intrauterine fetal death.

Keywords: protein C, protein S, thrombophelia, pregnancy, protein Z

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Authors: Natan C. G. Silva, Carlos A. C. Girao Neto, Marcele M. S. Vasconcelos, Luciana R. B. Goncalves, Maria Valderez P. Rocha

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Galactooligosaccharides (GOS) are sugars with prebiotic function that can be synthesized chemically or enzymatically, and this last one can be promoted by the action of β-galactosidases. In addition to favoring the transgalactosylation reaction to form GOS, these enzymes can also catalyze the hydrolysis of lactose. A highly studied type of GOS is lactulose because it presents therapeutic properties and is a health promoter. Among the different raw materials that can be used to produce lactulose, whey stands out as the main by-product of cheese manufacturing, and its discarded is harmful to the environment due to the residual lactose present. Therefore, its use is a promising alternative to solve this environmental problem. Thus, lactose from whey is hydrolyzed into glucose and galactose by β-galactosidases. However, in order to favor the transgalactosylation reaction, the medium must contain fructose, due this sugar reacts with galactose to produce lactulose. Then, the glucose-isomerase enzyme can be used for this purpose, since it promotes the isomerization of glucose into fructose. In this scenario, the aim of the present work was first to develop β-galactosidase biocatalysts of Kluyveromyces lactis and to apply it in the integrated reactions of hydrolysis, isomerization (with the glucose-isomerase from Streptomyces murinus) and transgalactosylation reaction, using whey as a substrate. The immobilization of β-galactosidase in chitosan previously functionalized with 0.8% glutaraldehyde was evaluated using different enzymatic loads (2, 5, 7, 10, and 12 mg/g). Subsequently, the hydrolysis and transgalactosylation reactions were studied and conducted at 50°C, 120 RPM for 20 minutes. In parallel, the isomerization of glucose into fructose was evaluated under conditions of 70°C, 750 RPM for 90 min. After, the integration of the three processes for the production of lactulose was investigated. Among the evaluated loads, 7 mg/g was chosen because the best activity of the derivative (44.3 U/g) was obtained, being this parameter determinant for the reaction stages. The other parameters of immobilization yield (87.58%) and recovered activity (46.47%) were also satisfactory compared to the other conditions. Regarding the integrated process, 94.96% of lactose was converted, achieving 37.56 g/L and 37.97 g/L of glucose and galactose, respectively. In the isomerization step, conversion of 38.40% of glucose was observed, obtaining a concentration of 12.47 g/L fructose. In the transgalactosylation reaction was produced 13.15 g/L lactulose after 5 min. However, in the integrated process, there was no formation of lactulose, but it was produced other GOS at the same time. The high galactose concentration in the medium probably favored the reaction of synthesis of these other GOS. Therefore, the integrated process proved feasible for possible production of prebiotics. In addition, this process can be economically viable due to the use of an industrial residue as a substrate, but it is necessary a more detailed investigation of the transgalactosilation reaction.

Keywords: beta-galactosidase, glucose-isomerase, galactooligosaccharides, lactulose, whey

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Authors: Alexander Hudson, Shaogang Gong

Abstract:

Structure determination is key to understanding protein function at a molecular level. Whilst significant advances have been made in predicting structure and function from amino acid sequence, researchers must still rely on expensive, time-consuming analytical methods to visualise detailed protein conformation. In this study, we demonstrate that it is possible to make accurate (≥80%) predictions of protein class and architecture from structures determined at low (>3A) resolution, using a deep convolutional neural network trained on high-resolution (≤3A) structures represented as 2D matrices. Thus, we provide proof of concept for high-speed, low-cost protein structure classification at low resolution, and a basis for extension to prediction of function. We investigate the impact of the input representation on classification performance, showing that side-chain information may not be necessary for fine-grained structure predictions. Finally, we confirm that high resolution, low-resolution and NMR-determined structures inhabit a common feature space, and thus provide a theoretical foundation for boosting with single-image super-resolution.

Keywords: transfer learning, protein distance maps, protein structure classification, neural networks

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Authors: Diana Karina Baigts Allende, Mariana Gonzalez Diaz, Luis Antonio Chel Guerrero, Mukthar Sandoval Peraza

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Poverty and food insecurity are still incident problems in the developing countries, where population´s diet is based on cereals which are lack in protein content. Nevertheless, during last years the use of native plants has been studied as an alternative source of protein in order to improve the nutritional intake. Chaya crop also called Spinach tree, is a prehispanic plant native from Central America and South of Mexico (Mayan culture), which has been especially valued due to its high nutritional content particularly protein and some medicinal properties. The aim of this work was to study the effect of protein isolation processing from Chaya leaf harvest in Yucatan, Mexico on its structure quality in order: i) to valorize the Chaya crop and ii) to produce low-cost and high-quality protein. Chaya leaf was extruded, clarified and recovered using: a) acid precipitation by decreasing the pH value until reach the isoelectric point (3.5) and b) thermal coagulation, by heating the protein solution at 80 °C during 30 min. Solubilized protein was re-dissolved in water and spray dried. The presence of Fraction I protein, known as RuBisCO (Rubilose-1,5-biphosfate carboxylase/oxygenase) was confirmed by gel electrophoresis (SDS-PAGE) where molecular weight bands of 55 KDa and 12 KDa were observed. The infrared spectrum showed changes in protein structure due to the isolation method. The use of high temperatures (thermal coagulation) highly decreased protein solubility in comparison to isoelectric precipitated protein, the nutritional properties according to amino acid profile was also disturbed, showing minor amounts of overall essential amino acids from 435.9 to 367.8 mg/g. Chaya protein isolate obtained by acid precipitation showed higher protein quality according to essential amino acid score compared to FAO recommendations, which could represent an important sustainable source of protein for human consumption.

Keywords: chaya leaf, nutritional properties, protein isolate, protein structure

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Authors: Alexandre Barbosa de Almeida, Telma Woerle de Lima Soares

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Protein structure prediction is a challenging task in the bioinformatics field. The biological function of all proteins majorly relies on the shape of their three-dimensional conformational structure, but less than 1% of all known proteins in the world have their structure solved. This work proposes a deep learning model to address this problem, attempting to predict some aspects of the protein conformations. Throughout a process of multiobjective dominance, a recurrent neural network was trained to abstract the particular bias of each individual multiobjective algorithm, generating a heuristic that could be useful to predict some of the relevant aspects of the three-dimensional conformation process formation, known as protein folding.

Keywords: Ab initio heuristic modeling, multiobjective optimization, protein structure prediction, recurrent neural network

Procedia PDF Downloads 177