• Asian-Australasian Journal of Animal Sciences
• /
• v.33 no.4
• /
• pp.556-567
• /
• 2020

Objective: To improve the utility of native grass resources as feed in China, we investigated the dynamics of protein and carbohydrate fractions among Inner Mongolian native grasses, during ensiling and the aerobic stage, using the Cornell Net Carbohydrate and Protein System. Methods: Silages were prepared without or with lactic acid bacteria (LAB) inoculant. We analyzed the protein and carbohydrate fractions and fermentation quality of silages at 0, 5, 15, 20, 30, and 60 d of ensiling, and the stability at 0.5, 2, 5, and 10 d during the aerobic stage. Results: Inner Mongolian native grass contained 10.8% crude protein (CP) and 3.6% water-soluble carbohydrates (WSC) on a dry matter basis. During ensiling, pH and CP and WSC content decreased (p<0.05), whereas lactic acid and ammonia nitrogen (N) content increased (p<0.05). Non-protein N (PA) content increased significantly, whereas rapidly degraded true protein (PB₁), intermediately degraded true protein (PB₂), total carbohydrate (CHO), sugars (CA), starch (CB₁), and degradable cell wall carbohydrate (CB₂) content decreased during ensiling (p<0.05). At 30 d of ensiling, control and LAB-treated silages were well preserved and had lower pH (<4.2) and ammonia-N content (<0.4 g/kg of fresh matter [FM]) and higher lactic acid content (>1.0% of FM). During the aerobic stage, CP, extract ether, WSC, lactic acid, acetic acid, PB₁, PB₂, true protein degraded slowly (PB₃), CHO, CA, CB₁, and CB₂ content decreased significantly in all silages, whereas pH, ammonia-N, PA, and bound true protein (PC) content increased significantly. Conclusion: Control and LAB-treated silages produced similar results in terms of fermentation quality, aerobic stability, and protein and carbohydrate fractions. Inner Mongolian native grass produced good silage, nutrients were preserved during ensiling and protein and carbohydrate losses largely occurred during the aerobic stage.

• Asian-Australasian Journal of Animal Sciences
• /
• v.32 no.10
• /
• pp.1521-1527
• /
• 2019

Objective: To study the contribution of plant enzyme and microbial activities on protein degradation in silage, this study evaluated the nitrogen transformation dynamics during ensiling of non- and irradiated alfalfa (Medicago sativa L.) and red clover (Trifolium pratense L.). Methods: Alfalfa and red clover silages were prepared and equally divided into two groups. One group was exposed to ${\gamma}$-irradiation at a recommended dosage (25 Gky). Therefore, four types of silages were produced: i) non-irradiated alfalfa silage; ii) irradiated alfalfa silage; iii) non-irradiated red clover silage; and iv) irradiated red clover silage. These silages were opened for fermentation quality and nitrogen components analyses after 1, 4, 8, and 30 days, respectively. Results: The ${\gamma}$-irradiation successfully suppressed microbial activity, indicated by high pH and no apparent increases in fermentation end products in irradiated silages. All nitrogen components, except for peptide-N, increased throughout the ensiling process. Proteolysis less occurred in red clover silages compared with alfalfa silages, indicated by smaller (p<0.05) increment in peptide-N and free amino acid N (FAA-N) during early stage of ensiling. The ${\gamma}$-irradiation treatment increased (p<0.05) peptide-N and FAA-N in alfalfa silage at day 1, whereas not in red clover silage; these two nitrogen components were higher (p<0.05) between day 4 and day 30 in non-irradiated silages than the irradiated silages. The ammonia nitrogen and non-protein nitrogen were highest in non-irradiated alfalfa silage and lowest in irradiated red clover silage after ensiling. Conclusion: The result of this study indicate that red clover and alfalfa are two forages varying in their nitrogen transformation patterns, especially during early stages of ensiling. Microbial activity plays a certain role in the proteolysis and seems little affected by the presence of polyphenol oxidase in red clover compared with alfalfaa.

• Bulletin of the Korean Chemical Society
• /
• v.33 no.8
• /
• pp.2508-2512
• /
• 2012

Cold shock proteins (CSPs) are a family of proteins induced at low temperatures. CSPs bind to single-stranded nucleic acids through the ribonucleoprotein 1 and 2 (RNP 1 and 2) binding motifs. CSPs play an essential role in cold adaptation by regulating transcription and translation via molecular chaperones. The solution nuclear magnetic resonance (NMR) or X-ray crystal structures of several CSPs from various microorganisms have been determined, but structural characteristics of psychrophilic CSPs have not been studied. Therefore, we optimized the purification process to obtain highly pure Lm-Csp and determined the three-dimensional structure model of Lm-Csp by comparative homology modeling using MODELLER on the basis of the solution NMR structure of Bs-CspB. Lm-Csp consists of a ${\beta}$-barrel structure, which includes antiparallel ${\beta}$ strands (G4-N10, F15-I18, V26-H29, A46-D50, and P58-Q64). The template protein, Bs-CspB, shares a similar ${\beta}$ sheet structure and an identical chain fold to Lm-Csp. However, the sheets in Lm-Csp were much shorter than those of Bs-CspB. The Lm-Csp side chains, E2 and R20 form a salt bridge, thus, stabilizing the Lm-Csp structure. To evaluate the contribution of this ionic interaction as well as that of the hydrophobic patch on protein stability, we investigated the secondary structures of wild type and mutant protein (W8, F15, and R20) of Lm-Csp using circular dichroism (CD) spectroscopy. The results showed that solvent-exposed aromatic side chains as well as residues participating in ionic interactions are very important for structural stability. Further studies on the three-dimensional structure and dynamics of Lm-Csp using NMR spectroscopy are required.

• Journal of Biomedical Engineering Research
• /
• v.14 no.4
• /
• pp.307-314
• /
• 1993

Plasma protein adsorption is the first event in the blood-material interaction and influenc- es subsequent platelet adhesion towards thlㅈombus formation. Thiㅈomboembolic events are strongly influenced by surface characteristics of materials and fluid dynamics inside the blood pump. In vitro flow visualizaion and an amimal experiment with the moving actuator type TAH were Performed in order to investigate fluid dynamic effects on the protein adsorption. The diffel'encl level, j of shear rate inside the ventricle Lvere determined by consid- ering the direction of the major opening of four healt valves in the implanted TAH and the visualized flow patterns as well. Each ventricle of the explanted TAH was sectionalized into 12 segments according to the shear rate level. The adsorbed protein on each segment was quantified using the ELISA method after soaking in 2% (wye)SDS/PBS for two days. Adsorbed protein layer thicknesses Itvere measured by the Immunogotd method under TEM. The SEM observation show that right ventricle (RV) , immobilized with albumin, displayed different degrees of platelet adhesion on each segment, whereas the left ventricle (LV), grafted by PEO-sulronate, indicated nearly , iame platelet adhesion behavior, regardless of shear rates. The surface concentrations of adsorbed proteins in the low shear rate region are hlghel'than those in the high region, which was confirmed statistically. A modified adsorption model of plasma protein onto polyurethane surface was suggested by considering the effect of the fluid dynamic characteristics.

• Proceedings of the Korean Society of Crop Science Conference
• /
• 2019.09a
• /
• pp.22-22
• /
• 2019

Green stem disorder (GSD) of soybean (Glycine max (L.) Merr.) is characterized by delayed senescence of stems with normal pod ripening and seed maturation (Hobbs, 2006). GSD complicates harvesting of soybeans by significantly increasing the difficulty in cutting the affected plants. There is also the potential for moisture in the stems to be scattered on the seed, reducing the grade and storability of the seed. Not only the cause of GSD is yet unknown, but also GSD cannot be evaluated until maturity, therefore the method to evaluate GSD in early growth stage with high sensitivity is necessary. In previous studies, it has been reported that vegetative storage protein (VSP) accumulates and the syndrome of GSD appears in soybean after depod treatment (Fischer, 1999). Soybean VSP is a storage protein which is abundant in young sink leaves and degraded during seed fill (Wittenbach, 1982). Hence, we have established a system to quantify VSP of high sensitivity by using standard protein made by genetically transformed E. coli and specific antibody against VSP, and studied the relationship between VSP and GSD, by depod experiment and drought/excess wet experiments. The result of depod experiment with the cultivar 'Yukihomare' was the same with the previous studies, VSP accumulated much more than control and the syndrome of GSD appeared in soybean in depod treatment. Drought and excess wet had different impact on GSD. Excess wet caused GSD of the cultivar 'Tachinagaha (GSD susceptible)', while drought caused a little syndrome of GSD in the cultivar 'Touhoku 129 (GSD resistant)'. The accumulation of VSP differed between the two cultivars over time. In conclusion, the accumulation of VSP came along with the emergence of GSD. Different cultivars showed different response to drought and excess wet. In the future, it is expected that the dynamics of VSP will be elucidated in detail, leading to the development of early diagnosis technology for green stem disorder and the elucidation of mechanism of soybean GSD.

• Genomics & Informatics
• /
• v.20 no.3
• /
• pp.33.1-33.17
• /
• 2022

Nervous necrosis virus (NNV) is a deadly infectious disease that affects several fish species. It has been found that the NNV utilizes grouper heat shock cognate protein 70 (GHSC70) to enter the host cell. Thus, blocking the virus entry by targeting the responsible protein can protect the fishes from disease. The main objective of the study was to evaluate the inhibitory potentiality of 70 compounds of Azadirachta indica (Neem plant) which has been reported to show potential antiviral activity against various pathogens, but activity against the NNV has not yet been reported. The binding affinity of 70 compounds was calculated against the GHSC70 with the docking and molecular dynamics (MD) simulation approaches. Both the docking and MD methods predict 4 (PubChem CID: 14492795, 10134, 5280863, and 11119228) inhibitory compounds that bind strongly with the GHSC70 protein with a binding affinity of -9.7, -9.5, -9.1, and -9.0 kcal/mol, respectively. Also, the ADMET (absorption, distribution, metabolism, excretion, and toxicity) properties of the compounds confirmed the drug-likeness properties. As a result of the investigation, it may be inferred that Neem plant compounds may act as significant inhibitors of viral entry into the host cell. More in-vitro testing is needed to establish their effectiveness.

• Genomics & Informatics
• /
• v.21 no.3
• /
• pp.42.1-42.23
• /
• 2023

Mycobacterium tuberculosis (Mtb) is the causative agent of tuberculosis, one of the most deadly infections in humans. The emergence of multidrug-resistant and extensively drug-resistant Mtb strains presents a global challenge. Mtb has shown resistance to many frontline antibiotics, including rifampicin, kanamycin, isoniazid, and capreomycin. The only licensed vaccine, Bacille Calmette-Guerin, does not efficiently protect against adult pulmonary tuberculosis. Therefore, it is urgently necessary to develop new vaccines to prevent infections caused by these strains. We used a subtractive proteomics approach on 23 virulent Mtb strains and identified a conserved membrane protein (MmpL4, NP_214964.1) as both a potential drug target and vaccine candidate. MmpL4 is a non-homologous essential protein in the host and is involved in the pathogen-specific pathway. Furthermore, MmpL4 shows no homology with anti-targets and has limited homology to human gut microflora, potentially reducing the likelihood of adverse effects and cross-reactivity if therapeutics specific to this protein are developed. Subsequently, we constructed a highly soluble, safe, antigenic, and stable multi-subunit vaccine from the MmpL4 protein using immunoinformatics. Molecular dynamics simulations revealed the stability of the vaccine-bound Tolllike receptor-4 complex on a nanosecond scale, and immune simulations indicated strong primary and secondary immune responses in the host. Therefore, our study identifies a new target that could expedite the design of effective therapeutics, and the designed vaccine should be validated. Future directions include an extensive molecular interaction analysis, in silico cloning, wet-lab experiments, and evaluation and comparison of the designed candidate as both a DNA vaccine and protein vaccine.

• Bulletin of the Korean Chemical Society
• /
• v.33 no.3
• /
• pp.828-832
• /
• 2012

We performed replica exchange molecular dynamics (REMD) simulations of the tripzip2 peptide (betahairpin) using the GB implicit and TI3P explicit solvation models. By comparing the resulting free energy surfaces of these two solvation model, we found that the GB solvation model produced a distorted free energy map, but the explicit solvation model yielded a reasonable free energy landscape with a precise location of the native structure in its global free energy minimum state. Our result showed that in particular, the GB solvation model failed to describe the tryptophan packing of trpzip2, leading to a distorted free energy landscape. When the GB solvation model is replaced with the explicit solvation model, the distortion of free energy shape disappears with the native-like structure in the lowest free energy minimum state and the experimentally observed tryptophan packing is precisely recovered. This finding indicates that the main source of this problem is due to artifact of the GB solvation model. Therefore, further efforts to refine this model are needed for better predictions of various aromatic side chain packing forms in proteins.

• BMB Reports
• /
• v.43 no.2
• /
• pp.127-132
• /
• 2010

Phosphatidylinositol (3,4,5)-triphosphate ($PIP_3$) is a lipid second messenger that employs a wide range of downstream effector proteins for the regulation of cellular processes, including cell survival, polarization and proliferation. One of the most well characterized cytoplasmic targets of $PIP_3$, serine/threonine protein kinase B (PKB)/Akt, promotes cell survival by directly interacting with nucleophosmin (NPM)/B23, the nuclear target of $PIP_3$. Here, we report that nuclear $PIP_3$ competes with Akt to preferentially bind B23 in the nucleoplasm. Mutation of Arg23 and Arg25 in the PH domain of Akt prevents binding to $PIP_3$, but does not disrupt the Akt/B23 interaction. However, treatment with phosphatases PTEN or SHIP abrogates the association between Akt and B23, indicating that nuclear $PIP_3$ regulates the Akt/B23 interaction by controlling the concentration and subcellular dynamics of these two proteins.

• Molecules and Cells
• /
• v.25 no.1
• /
• pp.131-137
• /
• 2008

Crk-associated substrate (CAS) is a focal adhesion protein that is involved in integrin signaling and cell migration. CAS deficiency reduces the migration and spreading of cells, both of which are processes mediated by Rac activation. We examined the functions of v-Crk, the oncogene product of the CT10 virus p47gag-crk, which affects cell migration and spreading, membrane ruffling, and Rac activation in CAS-deficient mouse embryonic fibroblasts (CAS-/- MEFs). CAS-/- MEFs showed less spreading than did CAS+/+ MEFs, but spreading was recovered in mutant cells that expressed v-Crk (CAS-/-v-Crk MEF). We observed that the reduction in spreading was linked to the formation of membrane ruffles, which were accompanied by Rac activation. In CAS-/- MEFs, Rac activity was significantly reduced, and Rac was not localized to the membrane. In contrast, Rac was active and localized to the membrane in CAS-/-v-Crk MEFs. Lamellipodia protrusion and ruffle retraction velocities were both reduced in CAS-/- MEFs, but not in CAS-/-v-Crk MEFs. We also found that microinjection of anti-gag antibodies inhibited the migration of CAS-/-v-Crk MEFs. These findings indicate that v-Crk controls cell migration and membrane dynamics by activating Rac in CAS-deficient MEFs.