Showing papers on "Gel electrophoresis published in 2022"
TL;DR: In this paper , the authors investigated the effects of combined treatment of enzyme and ultrasound on the structure of soybean protein isolate (SPI) and the properties of SPI gel cross-linked with glutamine transaminase (TG).
Abstract: This study aimed to investigate the effects of combined treatment of enzyme and ultrasound on the structure of soybean protein isolate (SPI) and the properties of SPI gel cross-linked with glutamine transaminase (TG). The papain-hydrolyzed SPI with hydrolysis degrees of about 0%, 0.1%, 0.5%, and 1.0% was treated with ultrasound (300 W, 20 min) to obtain different modified proteins. The sodium dodecyl sulfate–polyacrylamide gel electrophoresis of SPI after combined treatment showed lower molecular weight, indicating dissociation of some subunits into smaller units. In addition, the protein structure extended, promoting the exposure of free sulfhydryl and hydrophobic groups. Compared with untreated SPI, the SPI gel prepared by the combined treatment formed a uniform and dense gel network, and its gel strength and water-holding capacity (WHC) significantly improved. The gel strength and WHC were higher in the SPI gel treated with a combination of ultrasound and enzyme than that treated with ultrasound or enzyme alone, exhibiting the synergistic effect of enzymatic and ultrasonic treatment. In conclusion, the combined treatment of enzyme and ultrasound might be an effective way of improving the structure and gel properties of SPI.
24 citations
TL;DR: Wang et al. as discussed by the authors extracted gelatins from tuna skins using acid (STG-A), enzyme (STg-E), and hot water (ST G-H) methods with yields of 65.30 ± 1.56, 39.83 ± 1,61, and 50.97 ± 1., respectively.
Abstract: For full use of Skipjack tuna (Katsuwonus pelamis) canning by-products, gelatins (STG) were extracted from tuna skins using acid (STG-A), enzyme (STG-E) and hot water (STG-H) methods with yields of 65.30 ± 1.56%, 39.83 ± 1.61%, and 50.97 ± 1.44%, respectively. Due to the high imino acid contents (228, 216, and 213 residues/1000 residues for STG-A, STG-E, and STG-H, respectively), STG-A showed the highest transparency and gel strength properties. Amino acid analysis, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), ultraviolet (UV) and Fourier transform infrared spectroscopy (FTIR) analysis confirmed that STG-A, STG-E, and STG-H kept the main structure of type I collagen, but enzyme and hot water extraction methods showed much stronger hydrolysis ability on α and β chains. Moreover, neutrase hydrolysate (STG-AH) of STG-A showed the highest 1,1-diphenyl-2-picrylhydrazyl radical (DPPH•) scavenging activity and the fraction STG-AH-І (<3.5 kDa) from STG-AH prepared by ultrafiltration could significantly protect human skin fibroblasts (HSFBs) against ultraviolet-A (UVA) injury. Furthermore, nineteen peptides with high antioxidant activity were purified from STG-AH-І and identified. These results suggested that STG-AH and its derived antioxidant peptides could serve as potential ingredients applied in health benefiting products for preventing UVA injury.
17 citations
TL;DR: In this article, the gel properties of myofibrillar proteins (MPs) from four meat sources (fish, beef, sheep, and pork) were compared, and the results provided useful information for developing mixed gel products with different gel strengths.
Abstract: In this study, the gel properties of myofibrillar proteins (MPs) from four meat sources (fish, beef, sheep, and pork) were compared. Oscillatory rheology measurements including temperature sweep, frequency sweep, and strain sweep were conducted to characterise the small and large deformation rheological properties of the MPs. In addition, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and scanning electron microscopy (SEM) were used to evaluate differences in the molecular weight distribution as well as the microstructures in gel among different MPs. Frequency sweep measurements showed that all MP gels were weak gels. MPs extracted from pork exhibited the highest gel strength and most compact gel structure, whereas those from fish exhibited the lowest gel strength and loosest gel structure. In addition, the MP extracted from pork (PSM) had the highest content of myosin heavy chain (MHC) and actin. In conclusion, the MPs extracted from fish source and mammalian sources varied significantly in terms of rheological properties and microstructural characteristics. These results provided useful information for developing mixed gel products with different gel strengths.
16 citations
TL;DR: In this paper, the physical properties and protein conformation of shrimp surimi at 4°C storage were investigated for fresh shrimp (F0) and shrimp frozen for 4/8 months (F4/F8).
Abstract: Variations in the cold gelation of shrimp surimi at low temperatures significantly affect their gel properties and secondary processing. In this study, the physical properties and protein conformation of shrimp surimi at 4 °C storage were investigated for fresh shrimp (F0) and shrimp frozen for 4/8 months (F4/F8). Gel hardness and frequency tests indicated that F0 maintained a sol state, whereas F4 and F8 exhibited elastic gel characteristics, and a higher degree of crosslinking occurred in F8 during storage. Gel electrophoresis revealed that the myosin heavy chain underwent polymerization in F4 and F8 during cold storage. The Raman spectra showed that F4 and F8 exhibited lower α-helix and higher β-sheet contents after storage. The intensity of C–H bending peaks and the ratio of tyrosine doublet bands also decreased significantly in F4 and F8. The changes in the secondary and tertiary structures in F8 were more notable than those in F4. Scanning electron microscopy confirmed the formation of the most compact gel network in F8. These results helped clarify the gelation process of shrimp surimi prepared from fresh and frozen states at low temperatures and contributed to regulate the gelation properties of shrimp surimi products.
16 citations
TL;DR: In this article , the gel properties of myofibrillar proteins (MPs) from four meat sources (fish, beef, sheep, and pork) were compared, and the results provided useful information for developing mixed gel products with different gel strengths.
Abstract: In this study, the gel properties of myofibrillar proteins (MPs) from four meat sources (fish, beef, sheep, and pork) were compared. Oscillatory rheology measurements including temperature sweep, frequency sweep, and strain sweep were conducted to characterise the small and large deformation rheological properties of the MPs. In addition, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and scanning electron microscopy (SEM) were used to evaluate differences in the molecular weight distribution as well as the microstructures in gel among different MPs. Frequency sweep measurements showed that all MP gels were weak gels. MPs extracted from pork exhibited the highest gel strength and most compact gel structure, whereas those from fish exhibited the lowest gel strength and loosest gel structure. In addition, the MP extracted from pork (PSM) had the highest content of myosin heavy chain (MHC) and actin. In conclusion, the MPs extracted from fish source and mammalian sources varied significantly in terms of rheological properties and microstructural characteristics. These results provided useful information for developing mixed gel products with different gel strengths.
14 citations
01 Jan 2022
TL;DR: In this article , the physical properties and protein conformation of shrimp surimi at 4 °C storage were investigated for fresh shrimp (F0) and shrimp frozen for 4/8 months (F4/F8).
Abstract: Variations in the cold gelation of shrimp surimi at low temperatures significantly affect their gel properties and secondary processing. In this study, the physical properties and protein conformation of shrimp surimi at 4 °C storage were investigated for fresh shrimp (F0) and shrimp frozen for 4/8 months (F4/F8). Gel hardness and frequency tests indicated that F0 maintained a sol state, whereas F4 and F8 exhibited elastic gel characteristics, and a higher degree of crosslinking occurred in F8 during storage. Gel electrophoresis revealed that the myosin heavy chain underwent polymerization in F4 and F8 during cold storage. The Raman spectra showed that F4 and F8 exhibited lower α-helix and higher β-sheet contents after storage. The intensity of C–H bending peaks and the ratio of tyrosine doublet bands also decreased significantly in F4 and F8. The changes in the secondary and tertiary structures in F8 were more notable than those in F4. Scanning electron microscopy confirmed the formation of the most compact gel network in F8. These results helped clarify the gelation process of shrimp surimi prepared from fresh and frozen states at low temperatures and contributed to regulate the gelation properties of shrimp surimi products.
12 citations
TL;DR: In this article , the effects of solvent additives on proteins using agarose native gel electrophoresis were investigated using a set of experiments with bovine serum albumin (BSA) induced by heating.
12 citations
TL;DR: In this paper , the content and migration pattern of Gd and Gl protein isolates were quantitatively compared and the percentage of vicilins in total seed protein was higher in Gd (74.9%) than in Gl (63.4%).
Abstract: Glanded (Gd) cottonseed (Gossypium hirsutum L.) contains scattered gossypol glands. Glandless (Gl) cottonseed is a new type of seed containing only trace levels of gossypol. This work quantitatively compared the content and migration pattern of Gd and Gl protein isolates. Both protein samples were subjected to sodium dodecyl sulfate (SDS)‐gel electrophoresis, and the protein gel bands were separated into seven partitions for peptide mass spectroscopic analysis. While multiple peptide fragments (isoformers) of vicilin and legumin proteins were present in both samples, the percentage of vicilins in total seed protein was higher in Gd (74.9%) than in Gl (63.4%). In contrast, legumin proteins were more abundant in Gl (30.4%) than Gd (23.6%). Minor protein components such as lipid‐related oleosins and vicilin‐like antimicrobial peptides 2‐2 were also observed at a relatively higher incidence in Gl compared with Gd, potentially reflecting a need for increased protein‐related defense capability in the absence of gossypol against natural predators or adverse growth environment.
10 citations
TL;DR: In this article , the effect of substituents on the diiamine ligands over mode of interaction between Casiopeinas and DNA has been studied by gel electrophoresis, UV-vis and circular dichroism (CD) spectroscopic techniques.
8 citations
TL;DR: In this article , the authors characterize key parameters that affect the migration pattern of circular RNA in gel electrophoresis systems, which include gel type, electrophoreis time, sample buffer composition, and voltage.
8 citations
TL;DR: A new Western blotting method of native proteins from agarose-based gel electrophoresis using a buffer at pH 6.1 containing basic histidine and acidic 2-(N-morpholino)ethanesulfonic acid successfully provided native structures for a variety of proteins and macromolecular complexes.
Abstract: We have developed a new Western blotting method of native proteins from agarose-based gel electrophoresis using a buffer at pH 6.1 containing basic histidine and acidic 2-(N-morpholino)ethanesulfonic acid. This gel electrophoresis successfully provided native structures for a variety of proteins and macromolecular complexes. This paper is focused on the Western blotting of native protein bands separated on agarose gels. Two blotting methods from agarose gel to PVDF membrane are introduced here, one by contact (diffusion) blotting and another by electroblotting after pre-treating the agarose gels with SDS. The contact blotting resulted in the transfer of native GFP, native human plexin domain containing protein 2 (PLXDC2) and native SARS-CoV-2 spike protein, which were detected by conformation-specific antibodies generated in-house.
TL;DR: Evaluation of the results suggests that assessment of the relative quantity of amplicons by band brightness is precise enough even for post-PCR analysis steps requiring PCR product concentrations within a certain range to function properly.
Abstract: Agarose gel electrophoresis is a relatively easy to use method, commonly applied to evaluate PCR reaction success. Intercalating agents or dyes are used to visualize the amplified fragments. However, it is uncertain to what extent the brightness of bands is informative about the concentration of the amplicons. To more closely examine the suitability of agarose gel electrophoresis to assess PCR product yield, we quantified the brightness of bands on a gel and compared these data with the results from spectrophotometry, fluorometry and qPCR. Evaluation of the results suggests that assessment of the relative quantity of amplicons by band brightness is precise enough even for post-PCR analysis steps requiring PCR product concentrations within a certain range to function properly.
TL;DR: In this article , a protocol for vertical and horizontal formats of agarose native gel electrophoresis is described followed by different staining procedures, which can be applied to specific cases and the advantages or caveats of the present technology.
Abstract: Electrophoresis is one of the most important analytical technologies for characterization of macromolecules and their interactions. Among them, native gel electrophoresis is used to analyze the macromolecules in the native structure. It differs in principle and information from those obtained by sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE) or blue native polyacrylamide gel electrophoresis (BN-PAGE). SDS-PAGE is carried out in the presence of strong denaturant, SDS, while BN-PAGE is done in the presence of negatively charged dye, e.g., Coomassie brilliant blue, G-250. Here, we describe native gel electrophoresis using agarose gel and a buffer at pH 6.1 composed of histidine and 2-(N-morpholino) ethanesulfonic acid. First, a protocol for vertical and horizontal formats of agarose native gel electrophoresis is described followed by different staining procedures. Then, various examples obtained using the developed procedure will be shown to demonstrate how the technology can be applied to specific cases and the advantages or caveats of the present technology.
TL;DR: This report describes fragmentation patterns of bLF dosed vaginally in clinical trials or incubated ex vivo with VF, and finds evidence that exopeptidases trimmed their N- and C-termini.
Abstract: When bovine lactoferrin (bLF) contacts human vaginal fluid (VF) it is subjected to proteolytic degradation. This report describes fragmentation patterns of bLF dosed vaginally in clinical trials or incubated ex vivo with VF. A consensus pattern of fragments was observed in samples from different women. The 80 kDa bLF molecule is initially cleaved between its homologous 40 kDa domains, the N-lobe and C-lobe, and then degraded into sub-fragments and mixtures of small peptides. We characterized this fragmentation process by polyacrylamide gel electrophoresis, western blotting, chromatographic separation, and mass spectral sequence analysis. Common to most VF fragmentation patterns were large amounts of an N-lobe 37 kDa fragment and a C-lobe 43 kDa fragment resulting from a single cleavage following tyrosine 324. Both fragments possessed full sets of iron-ligand amino acids and retained iron-binding ability. In some VF samples, alternative forms of large fragments were found, which like the 37+43 kDa pair, totaled 80 kDa. These included 58+22 kDa, 18+62 kDa, and 16+64 kDa forms. In general, the smaller component was from the N-lobe and the larger from the C-lobe. The 18+62 kDa pair was absent in some VF samples but highly abundant in others. This variability suggests multiple endopeptidases are involved, with the 18 kDa fragment’s presence dependent upon the balance of enzymes. Further action of VF endopeptidases produced smaller peptide fragments, and we found evidence that exopeptidases trimmed their N- and C-termini. The 3.1 kDa antimicrobial peptide lactoferricin B was not detected. These studies were facilitated by a novel technique we developed: tricolor western blots, which enabled simultaneous visualization of N- and C-terminal epitopes.
TL;DR: In this paper , the protein content of granular cells and semigranular cells (SGCs) was compared by using two workflows: one-dimensional gel electrophoresis followed by LC-MS/MS and in-solution digestion of cell lysate.
Abstract: Semigranular cells (SGCs) and granular cells (GCs) are two dominant groups of circulating hemocytes in crayfish Cherax quadricarinatus. Molecular markers are required for the clear classification of the hemocytes and the research of their function and differentiation. In this study, we compared the protein content of GCs and SGCs by using two workflows: one-dimensional gel electrophoresis followed by LC-MS/MS and in-solution digestion of cell lysate followed by LC-MS/MS. Cell type-specific proteins were identified, and their expression in SGCs and GCs was further investigated by RT-PCR, Western blotting, and immunofluorescence analysis. Three molecular markers for GCs (peroxinectin, a mannose-binding protein, and prophenoloxidase-activating enzyme 2a) and three molecular markers for SGCs (a vitelline membrane outer layer protein I-like protein, a C-type lectin, and a peptidase) were identified. The application of some of the markers in Eriocheir sinensis was also analyzed. These molecular markers are useful tools for the research of crustaceans hemocytes.
TL;DR: Investigating cfDNA fragmentation patterns in cultured human bone cancer cells using increasingly sensitive electrophoresis assays revealed an exponential decay in the relative contribution of increasingly longer cfDNA populations, which suggests the involvement of a stochastic inter-nucleosomal DNA cleavage process.
Abstract: Unique bits of genetic, biological and pathological information occur in differently sized cell-free DNA (cfDNA) populations. This is a significant discovery, but much of the phenomenon remains to be explored. We investigated cfDNA fragmentation patterns in cultured human bone cancer (143B) cells using increasingly sensitive electrophoresis assays, including four automated microfluidic capillary electrophoresis assays from Agilent, i.e., DNA 1000, High Sensitivity DNA, dsDNA 915 and dsDNA 930, and an optimized manual agarose gel electrophoresis protocol. This comparison showed that (i) as the sensitivity and resolution of the sizing methods increase incrementally, additional nucleosomal multiples are revealed (hepta-nucleosomes were detectable with manual agarose gel electrophoresis), while the estimated size range of high molecular weight (HMW) cfDNA fragments narrow correspondingly; (ii) the cfDNA laddering pattern extends well beyond the 1–3 nucleosomal multiples detected by commonly used methods; and (iii) the modal size of HMW cfDNA populations is exaggerated due to the limited resolving power of electrophoresis, and instead consists of several poly-nucleosomal subpopulations that continue the series of DNA laddering. Furthermore, the most sensitive automated assay used in this study (Agilent dsDNA 930) revealed an exponential decay in the relative contribution of increasingly longer cfDNA populations. This power-law distribution suggests the involvement of a stochastic inter-nucleosomal DNA cleavage process, wherein shorter populations accumulate rapidly as they are fed by the degradation of all larger populations. This may explain why similar size profiles have historically been reported for cfDNA populations originating from different processes, such as apoptosis, necrosis, accidental cell lysis and purported active release. These results not only demonstrate the diversity of size profiles generated by different methods, but also highlight the importance of caution when drawing conclusions on the mechanisms that generate different cfDNA size populations, especially when only a single method is used for sizing.
TL;DR: In this paper , total-cell protein profiles of all the strains isolated were resolved via SDS-Polyacrylamide Gel Electrophoresis (PAGE) and more than 15 protein bands were found ranging from 15 kDa to 153 kDa in size.
TL;DR: It was concluded that catalysis of protein cross-linking is a general property of organophosphorus pesticides and pesticide metabolites.
Abstract: Exposure to organophosphorus pesticides (OP) can have chronic adverse effects that are independent of inhibition of acetylcholinesterase, the classic target for acute OP toxicity. In pure proteins, the organophosphorus pesticide chlorpyrifos oxon induces a cross-link between lysine and glutamate (or aspartate) with loss of water. Tubulin is particularly sensitive to OP-induced cross-linking. Our goal was to explore OP-induced cross-linking in a complex protein sample, MAP-rich tubulin from Sus scrofa and to test 8 OP for their capacity to promote isopeptide cross-linking. We treated 100 μg of MAP-rich tubulin with 100 μM chlorpyrifos, chlorpyrifos oxon, methamidophos, paraoxon, diazinon, diazoxon, monocrotophos, or dichlorvos. Each sample was separated using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and stained with Coomassie blue. Five gel slices (at about 30, 50, 150, and 300 kDa, and the top of the separating gel) were removed from the lanes for each of the eight OP samples and from untreated control lanes. These gel slices were subjected to in-gel trypsin digestion. MSMS fragmentation spectra of the tryptic peptides were examined for isopeptide cross-links. Sixteen spectra yielded convincing evidence for isopeptide cross-linked peptides. Ten were from the chlorpyrifos oxon reaction, 1 from dichlorvos, 1 from paraoxon, 1 from diazinon, and 3 from diazoxon. It was concluded that catalysis of protein cross-linking is a general property of organophosphorus pesticides and pesticide metabolites. Data are available via ProteomeXchange with identifier PXD034529.
TL;DR: In this paper , the levels of cerebrospinal fluid (CSF) apolipoprotein E (apoE) species in Alzheimer's disease (AD) patients were examined.
Abstract: The purpose of this study was to examine the levels of cerebrospinal fluid (CSF) apolipoprotein E (apoE) species in Alzheimer's disease (AD) patients.We analyzed two CSF cohorts of AD and control individuals expressing different APOE genotypes. Moreover, CSF samples from the TgF344-AD rat model were included. Samples were run in native- and SDS-PAGE under reducing or non-reducing conditions (with or without β-mercaptoethanol). Immunoprecipitation combined with mass spectrometry or western blotting analyses served to assess the identity of apoE complexes.In TgF344-AD rats expressing a unique apoE variant resembling human apoE4, a ~35-kDa apoE monomer was identified, increasing at 16.5 months compared with wild-types. In humans, apoE isoforms form disulfide-linked dimers in CSF, except apoE4, which lacks a cysteine residue. Thus, controls showed a decrease in the apoE dimer/monomer quotient in the APOE ε3/ε4 group compared with ε3/ε3 by native electrophoresis. A major contribution of dimers was found in APOE ε3/ε4 AD cases, and, unexpectedly, dimers were also found in ε4/ε4 AD cases. Under reducing conditions, two apoE monomeric glycoforms at 36 kDa and at 34 kDa were found in all human samples. In AD patients, the amount of the 34-kDa species increased, while the 36-kDa/34-kDa quotient was lower compared with controls. Interestingly, under reducing conditions, a ~100-kDa apoE complex, the identity of which was confirmed by mass spectrometry, also appeared in human AD individuals across all APOE genotypes, suggesting the occurrence of aberrantly resistant apoE aggregates. A second independent cohort of CSF samples validated these results.These results indicate that despite the increase in total apoE content the apoE protein is altered in AD CSF, suggesting that function may be compromised.
TL;DR: Based on the combined procedures, conformation-specific monoclonal antibodies against PLXDC2 and SARS-CoV-2 spike protein were discovered and green fluorescent protein showed functional state both on agarose gel and blotted membrane.
Abstract: In this study, we review the agarose native gel electrophoresis that separates proteins and macromolecular complexes in their native state and transfer of the separated proteins from the agarose gel to membranes by contact blotting which retains the native state of these structures. Green fluorescent protein showed functional state both on agarose gel and blotted membrane. Based on the combined procedures, we discovered conformation-specific monoclonal antibodies against PLXDC2 and SARS-CoV-2 spike protein.
TL;DR: In this article , a technique for detecting protein developed based on protein electrophoresis separation and antigen-antibody detection is described, and the specific expression level of protein is analyzed by immunological detection.
Abstract: Western blotting is a technique for detecting protein developed based on protein electrophoresis separation and antigen-antibody detection. It combines the high resolution of SDS polyacrylamide gel electrophoresis with the specificity of antigen-antibody reaction. The basic principle is that the protein components are separated by electrophoresis, then the protein on the gel after electrophoresis is transferred to the carrier membrane, and the area of the carrier membrane where protein is not adsorbed is blocked by a blocking reagent. Finally, the specific expression level of protein is analyzed by immunological detection. It overcomes the disadvantages of direct immunological analysis on polyacrylamide gel after electrophoresis greatly improves its resolution and sensitivity, and is widely used to detect the correctness of specific gene expression products or compare the relative variation of proteins’ expression.
TL;DR: A sequential synthesis route towards annulated imidazo[4,5-c]isoquinolines comprising a GBB-3CR, followed by an intramolecular imidoylative cyclisation is reported, revealing a stabilising effect on ds-DNA against strand-break inducing conditions.
Abstract: Herein we report the development of a sequential synthesis route towards annulated imidazo[4,5-c]isoquinolines comprising a GBB-3CR, followed by an intramolecular imidoylative cyclisation. X-Ray crystallography revealed a flat 3D structure of the obtained polyheterocycles. Thus, we evaluated their interactions with double-stranded DNA by establishing a pUC-19 plasmid-based gel electrophoresis mobility shift assay, revealing a stabilising effect on ds-DNA against strand-break inducing conditions.
TL;DR: In this paper , three camera modules (V1.3 with and without filter, as well as NoIR) were assessed for their suitability in imaging fluorescent DNA following agarose gel electrophoresis.
Abstract: Low-cost analytical solutions built around microcomputers like the Raspberry Pi help to facilitate laboratory investigations in resource limited venues. Here, three camera modules (V1.3 with and without filter, as well as NoIR) that work with this microcomputer were assessed for their suitability in imaging fluorescent DNA following agarose gel electrophoresis. Evaluation of their utility was based on signal-to-noise (SNR) and noise variance metrics that were developed. Experiments conducted with samples were subjected to Polymerase Chain Reaction (PCR), and the amplified products were separated using gel electrophoresis and stained with Midori green. Image analysis revealed the NoIR camera performed the best with SNR and noise variance values of 21.7 and 0.222 respectively. In experiments conducted using UV LED lighting to simulate ethidium bromide (EtBr) excitation, the NoIR and V1.3 with filter removed cameras showed comparable SNR values.
TL;DR: In this paper , the effects of different levels (0, 1, 3, and 5, w/w) of cricket protein powder (CP) and soy protein isolate (SPI) on the gel properties of mackerel surimi were investigated.
Abstract: This work comparatively investigated the effects of different levels (0, 1, 3, and 5%, w/w) of cricket protein powder (CP) and soy protein isolate (SPI) on the gel properties of mackerel surimi. Both SPI and CP enhanced the rheological properties of surimi pastes during heating, as indicated by the increase in G′ and G″ and the decrease in tan δ. With increasing SPI content, the proteolytic inhibition, gel properties, water-holding capacity, and textural profiles of surimi gel were markedly enhanced. Molecular driving-force results showed that SPI markedly promoted the hydrophobic interaction, while disulfide bonds were dominant in CP-added gel. However, the whiteness of surimi gels tended to decrease with the increased levels of both additives, in particular CP. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) demonstrated that SPI hindered the polymerization of myosin heavy chain while CP participated in the formation of non-disulfide covalent bonds with actin. Fourier transform infrared (FTIR) spectra indicated that CP and SPI did not influence the secondary structure of proteins in surimi. Scanning electron microscopy (SEM) demonstrated that CP or SPI induced the myofibrillar protein to form smoother and compact gel network structures. Overall acceptability of the mackerel surimi gel can be improved by the incorporation of 5% SPI while CP had a negative impact on several parameters. However, CP showed the remarkable ability to prevent the lipid oxidation of the gel after storage at 4 °C for 7 days. Overall, both SPI and CP demonstrated positive impacts on the gelling characteristics of mackerel surimi; however, SPI was more advantageous than CP in terms of the gel-strengthening effect and sensory qualities. This study offered a potential use for plant and insect proteins as functional and nutritional ingredients for the production of dark-fleshed fish surimi.
TL;DR: This method fractionates RNA by electrophoresis through an agarose gel containing 2.2 m formaldehyde to denatured samples of RNA by treatment with formamide and separated through agaroses gels containing formaldehyde.
Abstract: Samples of RNA may be denatured by treatment with formamide and separated by electrophoresis through agarose gels containing formaldehyde. In this method, RNA is fractionated by electrophoresis through an agarose gel containing 2.2 m formaldehyde.
TL;DR: In this article , the effects of greening process on the functional and structural properties of garlic protein were investigated, and a proteomic strategy was applied to analyze the changes of protein compositions as well as their activities.
TL;DR: In this article , an attempt was made to specifically stain unfolded proteins on agarose native gels, using bovine serum albumin (BSA), which was shown that staining did not depend on whether BSA was thermally unfolded in the presence of SYPRO Orange or stained after electrophoresis.
TL;DR: Ultrasensitive, quantitative Clostridioides difficile stool toxin measurement demonstrated significantly higher concentrations of toxins A and B in patients infected with the NAP-1/027 strain compared to patientsinfected with other strains.
Abstract: Ultrasensitive, quantitative Clostridioides difficile stool toxin measurement demonstrated significantly higher concentrations of toxins A and B in patients infected with the NAP-1/027 strain compared to patients infected with other strains. These data provide in vivo confirmation of the in vitro association between NAP-1/027 strain type and elevated toxin production.
TL;DR: The recently developed Phos-tag diagonal electrophoresis has been able to solve the problem of comparing the electrophoretic mobility of the proteins in the different gels used, and can indicate the SDS-PAGE and PhOS-tag SDS -PAGE patterns on a single gel; therefore, phosphorylated proteins can be distinguished easily from non-phosphorylation proteins.
Abstract: ABSTRACT Introduction Phosphate-binding tag (Phos-tag) sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) is an important development capable of analyzing the phosphorylation state of proteins. Conventionally, proteins were separated via SDS-PAGE and Phos-tag SDS-PAGE that use different gels to identify phosphorylated proteins. However, it was often difficult to compare the electrophoretic mobility of the proteins in the different gels used. The recently developed Phos-tag diagonal electrophoresis has been able to solve this problem. It can indicate the SDS-PAGE and Phos-tag SDS-PAGE patterns on a single gel; therefore, phosphorylated proteins can be distinguished easily from non-phosphorylated proteins. Areas covered This review assesses the importance of Phos-tag electrophoresis, which enables the analysis of protein phosphorylation states, in the field of proteomics. Additionally, this review describes the significance and actual experimental technique of Phos-tag diagonal electrophoresis, which was recently developed to overcome the drawbacks of Phos-tag SDS-PAGE. Expert opinion Although shotgun analysis of proteins allows detecting many phosphorylation sites, it is challenging to clarify the differences in the phosphorylation states of protein molecules using this technique. Therefore, Phos-tag SDS-PAGE is frequently used to determine the phosphorylation state of proteins. This technique has become more powerful with the recent development of Phos-tag diagonal electrophoresis. Abbreviations: BIS, N,N’-methylenebis(acrylamide); CBB, Coomassie brilliant blue R250; ESI, electrospray ionization; hnRNP, heterogeneous ribonucleoprotein K; LTQ–Orbitrap, Linear trap quadrupole–Orbitrap; LC, liquid chromatography; MS, mass spectrometry; MALDI, matrix-assisted laser desorption ionization; Phos-tag, phosphate-binding tag [1,3-bis [bis (pyridine-2-ylmethyl) amino] propane-2-olate]; SDS-PAGE, sodium dodecyl sulfate-polyacrylamide gel electrophoresis; TOF, time of flight; 2D-DIGE, fluorescence-labeled two-dimensional difference gel electrophoresis; 2-DE, two-dimensional gel electrophoresis
TL;DR: In this paper , the structural proteins of the Decapod iridescent virus 1 (DIV1) were analyzed by mass spectrometry and 30 virus-encoded proteins were identified.