scholarly journals Study on Effect of Extraction Techniques and Seed Coat on Proteomic Distribution and Cheese Production from Soybean Milk

Molecules ◽  
2020 ◽  
Vol 25 (14) ◽  
pp. 3237
Author(s):  
Nadia Al-Saedi ◽  
Manjree Agarwal ◽  
Wujun Ma ◽  
Shahidul Islam ◽  
Yonglin Ren
Keyword(s):  
Protein Extraction ◽  
Consumer Preferences ◽  
Polyacrylamide Gel Electrophoresis ◽  
Separation Techniques ◽  
Soybean Milk ◽  
Allergenic Proteins ◽  
Reference Map ◽  
Whole Seed ◽  
Α Subunits ◽  
Cheese Production

Soybean-based food products are a major source of protein. In the present study, proteins in soybean milk from seeds of the cultivar Bunya (Glycine max) were extracted using the cheesecloth and the centrifuge methods. The milk was produced through mechanical crushing of both whole and split seeds in water. Following separation by either the cheesecloth or centrifuge, proteins were isolated from the soybean milk by using thiourea/urea solubilisation and then separated them using two-dimensional polyacrylamide gel electrophoresis. The isolated proteins were identified by mass spectrometry. A total of 97 spots were identified including 49 that displayed different abundances. Of the two separation techniques, centrifuge separation gave higher protein extraction and more intense protein spots than cheesecloth separation. Eleven of the β-subunits of β-conglycinin, three of the α-subunits of β-conglycinin, and four of the mutant glycinin showed different levels of abundances between separation techniques, which might be related to subsequent cheese quality. Notably, split-seed soybean milk has less allergenic proteins with four α-subunits of β-conglycinin compared to whole-seed milk with eight of those proteins. The sensory evaluation showed that the cheese produced from split-soybean milk received higher consumer preferences compared to that of whole seed, which could be explained by their proteomic differences. The demonstrated reference map for whole and split-seed soybean milk could be further utilized in the research related to soybean cheesemaking.

The reaction between hemoglobin α-subunit and haptoglobin

1976 ◽  
Vol 54 (11) ◽  
pp. 1011-1015 ◽  
Author(s):  
Frank A. Terpstra ◽  
David B. Smith
Keyword(s):  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Gel Filtration ◽  
Polyacrylamide Gel ◽  
Human Hemoglobin ◽  
Gel Filtration Chromatography ◽  
Separation Techniques ◽  
Α Subunit ◽  
Α Subunits

The interaction between human hemoglobin α-subunit and porcine haptoglobin was investigated by polyacrylamide gel electrophoresis, gel filtration chromatography, sedimentation through excess α-subunit and gel filtration in an α-subunit-containing medium. No interaction was detected by the first two methods indicating dissociation of the complex during the application of these separation techniques. The latter two methods, in which the complex is studied in a medium of excess subunits, showed that haptoglobin became saturated with the binding of two α-subunits.

scholarly journals Development of a Listeria monocytogenes EGDe Partial Proteome Reference Map and Comparison with the Protein Profiles of Food Isolates

2003 ◽  
Vol 69 (6) ◽  
pp. 3368-3376 ◽  
Author(s):  
Manilduth Ramnath ◽  
K. Bj�rn Rechinger ◽  
Lothar J�nsch ◽  
John W. Hastings ◽  
Susanne Kn�chel ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Total Protein ◽  
Protein Extraction ◽  
Polyacrylamide Gel Electrophoresis ◽  
Model Organism ◽  
Pcr Amplification ◽  
Major Protein ◽  
Starting Point ◽  
Serotype 1 ◽  
Reference Map

ABSTRACT A partially annotated proteome reference map of the food pathogen Listeria monocytogenes was developed for exponentially growing cells under standardized, optimal conditions by using the sequenced strain EGDe (serotype 1/2a) as a model organism. The map was developed by using a reproducible total protein extraction and two-dimensional (2-D) polyacrylamide gel electrophoresis analysis procedure, and it contained 33 identified proteins representing the four main protein functional classes. In order to facilitate analysis of membrane proteins, a protein compartmentalization procedure was assessed. The method used provided partial fractionation of membrane and cytosolic proteins. The total protein 2-D profiles of three serotype 1/2a strains and one serotype 1/2b strain isolated from food were compared to the L. monocytogenes EGDe proteome. An average of 13% of the major protein spots in the food strain proteomes were not matched in the strain EGDe proteome. The variation was greater for the less intense spots, and on average 28% of these spots were not matched. Two of the proteins identified in L. monocytogenes EGDe were missing in one or more of the food isolates. These two proteins were proteins involved in the main glycolytic pathway and in metabolism of coenzymes and prosthetic groups. The two corresponding genes were found by PCR amplification to be present in the four food isolates. Our results show that the L. monocytogenes EGDe reference map is a valuable starting point for analyses of strains having various origins and could be useful for analyzing the proteomes of different isolates of this pathogen.

scholarly journals A comparative study on the extraction effects of common agents on collagen-based binders in mural paintings

2021 ◽  
Vol 9 (1) ◽  
Author(s):  
Jianghao Du ◽  
Zhanyun Zhu ◽  
Junchang Yang ◽  
Jia Wang ◽  
Xiaotong Jiang
Keyword(s):  
Protein Structure ◽  
Comparative Study ◽  
Protein Extraction ◽  
Guanidine Hydrochloride ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Extraction Process ◽  
Ultraviolet Absorption Spectrum ◽  
Mural Paintings ◽  
The Impact

AbstractIn this paper, a comparative study was conducted on the extraction effects of six agents for collagen-based mural painting binders. These agents were used to extract the residual proteins in the non-aged and thermal aged samples. The protein extraction efficiencies of different extracting agents were quantitatively determined by bicinchoninic acid (BCA) method, and then processed by multivariate analysis of variance (MANOVA). The impact of the extraction process on the protein structure was characterized by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), ultraviolet absorption spectrum (UV) and circular dichroism (CD). The results showed that, for both non-aged and aged samples, the extraction efficiency of 2 M guanidine hydrochloride (GuHCl) was significantly higher than the other five agents, with less damage to the protein structure during the extraction process.

scholarly journals Evaluation of effective protein extraction procedure to profile petiole of Moringa oleifera

2020 ◽  
Vol 7 (2) ◽  
pp. 214
Author(s):  
Zetty Amirah Zulkifli ◽  
Zaidah Rahmat
Keyword(s):  
Moringa Oleifera ◽  
Protein Extraction ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Extraction Procedure ◽  
Electrophoretic Pattern ◽  
Antioxidant Property ◽  
Extraction Methods ◽  
Protein Profiling ◽  
Sds Page

Moringa oleifera is widely known as multipurpose tree since all of its parts confer multiple functions. The leaf is highly favourable among consumers while the petiole is mostly wasted. There are numerous studies on the flavonoid and antioxidant property of the stem and twig. However, study on the petiole has never been done. There-upon, this study was conducted to develop protein profiling of the petiole. In this study, 6 different protein extraction methods were tested on the fresh petiole before its protein quantity and quality were checked via Bradford assay and Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis (SDS-PAGE) respectively. The in-solution digestion was then done prior to LC-MS/MS analysis. The protein electrophoretic pattern from the SDS-PAGE proves that method 6 using Tris HCl buffer with incorporation of dithiothreitol (DTT) and phenylmethylsulfonyl fluoride (PMSF) confers the best quality of protein. It produced the highest number of visible individual bands compared to other methods. Meanwhile, 93 proteins were successfully identified via LCMS analysis where the protein, signal response and carbohydrate metabolism categories confer the highest percentage. High quality and content of the protein extracted from the petiole including the antioxidant, anticancer and antidiabetic protein identified suggested that consuming this part of the plant could enhance nutrients of human body.

Protein extraction from activated sludge

2006 ◽  
Vol 54 (1) ◽  
pp. 175-181 ◽  
Author(s):  
M. Denecke
Keyword(s):  
Activated Sludge ◽  
Crude Extract ◽  
Extracellular Polymeric Substances ◽  
Protein Extraction ◽  
Polyacrylamide Gel Electrophoresis ◽  
Bacterial Cells ◽  
Uronic Acids ◽  
Isolated Cells ◽  
Humic Compounds ◽  
Immunoblot Technique

Two methods for the separation of protein originating from activated sludge were compared. In one method, the total protein was isolated out of the activated sludge (crude extract). These samples included all dissolved proteins originating from the bacterial cells and biofilm made up of extracellular polymeric substances (EPS). Every time polyacrylamide gel electrophoresis (PAGE) was done, the protein bands from samples of crude extract were covered by polymeric substances including carbohydrates, uronic acids or humic compounds. Using the immunoblot technique it was possible to demonstrate the presence of the heat shock protein HSP70 in crude extracts of activated sludge. The comparison of protein fingerprints required that clear and distinct bands appear on the PAGE analysis. To this end, a procedure to separates bacterial cells from the EPS was developed. Bacterial cells were separated by incubation with EDTA and subsequent filtration. The isolated cells were directly incubated in a sample buffer.

scholarly journals Differential Proteome Analysis of the Preeclamptic Placenta Using Optimized Protein Extraction

2010 ◽  
Vol 2010 ◽  
pp. 1-9 ◽  
Author(s):  
Magnus Centlow ◽  
Stefan R. Hansson ◽  
Charlotte Welinder
Keyword(s):  
Signal Transduction ◽  
Gel Electrophoresis ◽  
Protein Separation ◽  
Protein Extraction ◽  
Polyacrylamide Gel Electrophoresis ◽  
Proteome Analysis ◽  
Apolipoprotein A1 ◽  
Two Dimensional ◽  
2D Page ◽  
Nutritional Effect

The human placenta is a difficult tissue to work with using proteomic technology since it contains large amounts of lipids and glycogen. Both lipids and glycogen are known to interfere with the first step in the two-dimensional polyacrylamide gel electrophoresis (2D-PAGE), the isoelectric focusing. In order to gain the best possible protein separation on 2D-PAGE, an optimized sample preparation protocol for placental proteins was developed. Two different buffers, urea/CHAPS and Hepes, were used for solubilization in combination with six different precipitation methods. The removal of glycogen from the samples by centrifugation was crucial for the final proteome maps. Solubilization with urea/CHAPS in combination with dichloromethane/methanol or acidified acetone proved to be the best precipitation procedures. When applied to clinical placenta samples apolipoprotein A1 was found to be accumulated in the preeclamptic placenta, where it may either have a nutritional effect or act as a modifier of signal transduction.

COMPARISON OF ALLERGENIC PROTEINS OF SEA SNAIL (CERITHIDEA OBTUSA) AND FRESHWATER SNAIL (POMACEA CANALICULATA)

2016 ◽  
Vol 78 (11) ◽  
Author(s):  
Rosmilah Misnan ◽  
Norazlin Salahudin Abdul Aziz ◽  
Zailatul Hani Mohd Yadzir ◽  
Noormalin Abdullah ◽  
Faizal Bakhtiar ◽  
...  
Keyword(s):  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Freshwater Snail ◽  
Protein Profiling ◽  
Protein Fractions ◽  
Pomacea Canaliculata ◽  
Market Demand ◽  
Allergenic Proteins ◽  
Major Allergens ◽  
Minor Allergens

In Malaysian and certain Asian countries, snail has high market demand and popular to the local people as food. However, snail is also frequently reported as one of the worst food allergens, dominated by severe symptoms such as asthma and anaphylactic shock. Thus, the aims of this study is to determine the allergenicity of two species of edible snails; the local sea snail, Cerithidea obtusa and the freshwater snail Pomacea canaliculata. Snail extracts were prepared from the snail flesh and analyzed using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) to determine their protein profiling. Allergenic proteins were detected by immunoblotting test using sera from 10 snail-allergic patients. The snails contain 31 to 34 protein fractions between 11 to >250 kDa. The prominent bands were seen at 33, 42, 74 and 250 kDa. Immunoblotting detected 15 and 16 allergenic proteins in C. obtusa and P. canaliculata, respectively. Three protein fractions at 30, 33 and 42 kDa were identified as the major allergens of C. obtusa, while six major allergens at 30, 33, 42, 74, 124 and 218 kDa were detected in P. canaliculata. Various minor allergens were also detected in both snails. This study indicated that numerous proteins of C. obtusa and P. canaliculata were allergenic. Thus, combined allergen extracts of both snails are essential to be included in diagnosis of snail allergy among local allergic patients.  

scholarly journals Ralstonia eutropha H16 Flagellation Changes According to Nutrient Supply and State of Poly(3-Hydroxybutyrate) Accumulation

2008 ◽  
Vol 74 (14) ◽  
pp. 4477-4490 ◽  
Author(s):  
Matthias Raberg ◽  
Frank Reinecke ◽  
Rudolf Reichelt ◽  
Ursula Malkus ◽  
Simone König ◽  
...  
Keyword(s):  
Stationary Phase ◽  
Growth Phase ◽  
Exponential Growth ◽  
Time Course ◽  
Ralstonia Eutropha ◽  
Protein Extraction ◽  
Polyacrylamide Gel Electrophoresis ◽  
Exponential Growth Phase ◽  
Electron Microscopic ◽  
2D Page

ABSTRACT Two-dimensional polyacrylamide gel electrophoresis (2D PAGE), in combination with matrix-assisted laser desorption ionization-time of flight analysis, and the recently revealed genome sequence of Ralstonia eutropha H16 were employed to detect and identify proteins that are differentially expressed during different phases of poly(3-hydroxybutyric acid) (PHB) metabolism. For this, a modified protein extraction protocol applicable to PHB-harboring cells was developed to enable 2D PAGE-based proteome analysis of such cells. Subsequently, samples from (i) the exponential growth phase, (ii) the stationary growth phase permissive for PHB biosynthesis, and (iii) a phase permissive for PHB mobilization were analyzed. Among several proteins exhibiting quantitative changes during the time course of a cultivation experiment, flagellin, which is the main protein of bacterial flagella, was identified. Initial investigations that report on changes of flagellation for R. eutropha were done, but 2D PAGE and electron microscopic examinations of cells revealed clear evidence that R. eutropha exhibited further significant changes in flagellation depending on the life cycle, nutritional supply, and, in particular, PHB metabolism. The results of our study suggest that R. eutropha is strongly flagellated in the exponential growth phase and loses a certain number of flagella in transition to the stationary phase. In the stationary phase under conditions permissive for PHB biosynthesis, flagellation of cells admittedly stagnated. However, under conditions permissive for intracellular PHB mobilization after a nitrogen source was added to cells that are carbon deprived but with full PHB accumulation, flagella are lost. This might be due to a degradation of flagella; at least, the cells stopped flagellin synthesis while normal degradation continued. In contrast, under nutrient limitation or the loss of phasins, cells retained their flagella.

scholarly journals The effects of camel chymosin and Withania coagulans extract on camel and bovine milk cheeses

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Mustapha Mbye ◽  
Huda Mohamed ◽  
Abdul Raziq ◽  
Afaf Kamal-Eldin
Keyword(s):  
Incubation Temperature ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Bovine Milk ◽  
Enzyme Concentration ◽  
Withania Coagulans ◽  
Sds Page ◽  
Cheese Production ◽  
Endogenous Enzymes

AbstractWithania coagulans (W. coagulans) extract and camel chymosin have aspartic protease capable of coagulating milk for cheese production. This study investigated the quality of camel and bovine milk cheeses coagulated using Withania extracts, came chymosin, and their mixture in two experiments. In Experiment (1), a factorial design with four factors (W. coagulans, camel chymosin, incubation time, and incubation temperature) was performed. The effect of these factors on cheese’s yield and hardness were assessed. An enzyme concentration corresponding to a 36 µg/L of milk of W. coagulans, 50 IMCU/L of camel chymosin, holding time of 4 h, and incubation temperature of 60 °C provided the optimal textural hardness for both camel and bovine milk cheeses. Seven treatments were analyzed in experiment (2) were analyzed for physicochemical properties, yield, and sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGEitation). The results showed that pure Withania extract exhibited the lower coagulating effect resulting in cheeses with low yield, hardness, fat, protein, and total solids. The SDS-PAGE electropherograms of camel cheese showed several low molecular weight bands as compared to bovine cheese. This phenomenon is due to excessive proteolysis in camel cheese, which we believed is caused by the presence of endogenous enzymes.

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