enzymatic activity
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2023 ◽  
Vol 83 ◽  
Author(s):  
E. Ibáñez-Arancibia ◽  
J. G. Farías ◽  
I. Valdebenito

Abstract The cold storage of milt implies potentials alterations in its quality because the storage generates as main process, free radicals that produce spermatozoa membrane lipids damage with the consequent motility and fertilising capacity disruptions. To decrease the damage generated by free radicals the cells have antioxidant defences (proteins, enzymes, and low molecular weight substances). The objective of the present study evaluated the time storage effect and different antioxidants prepared in spermatic diluents on sperm viability of O. mykiss milt stored at 4°C. The two-way ANOVA denoted that the time storage and antioxidant influence have significant effects separated or combined on viability parameters (sperm motility and viability, proteins concentrations and superoxide dismutase enzymatic activity in seminal plasma). In contrast, only the storage time affected the fertilising capacity and catalase enzymatic activity in seminal plasma. The resulting analysis can conclude that the antioxidant presence improves the viability of cold stored milt, especially the transport conditions and the antioxidants allow the fecundity despite motility decrease.


Cells ◽  
2022 ◽  
Vol 11 (2) ◽  
pp. 298
Author(s):  
José Antonio de Mera-Rodríguez ◽  
Guadalupe Álvarez-Hernán ◽  
Yolanda Gañán ◽  
Ana Santos-Almeida ◽  
Gervasio Martín-Partido ◽  
...  

The histochemical detection of β-galactosidase enzymatic activity at pH 6.0 (β-gal-pH6) is a widely used biomarker of cellular senescence in aging tissues. This histochemical assay also detects the presence of programmed cell senescence during specific time windows in degenerating structures of vertebrate embryos. However, it has recently been shown that this enzymatic activity is also enhanced in subpopulations of differentiating neurons in the developing central nervous system in vertebrates. The present study addressed the histochemical detection of β-gal-pH6 enzymatic activity in the developing postnatal olfactory epithelium in the mouse. This activity was detected in the intermediate layer of the olfactory epithelium. As development progressed, the band of β-gal-pH6 labeling in this layer increased in width. Immunohistochemistry and lectin histochemistry showed the β-gal-pH6 staining to be strongly correlated with the immunolabeling of the olfactory marker protein (OMP) that identifies mature olfactory sensory neurons. The cell somata of a subpopulation of differentiated olfactory neurons that were recognized with the Dolichos biflorus agglutinin (DBA) were always located inside this band of β-gal-pH6 staining. However, the β-gal-pH6 histochemical signal was always absent from the apical region where the cytokeratin-8 positive supporting cells were located. Furthermore, no β-gal-pH6 staining was found in the basal region of the olfactory epithelium where PCNA/pHisH3 immunoreactive proliferating progenitor cells, GAP43 positive immature neurons, and cytokeratin-5 positive horizontal basal cells were located. Therefore, β-gal-pH6 seems to be linked to neuronal differentiation and cannot be regarded as a biomarker of cellular senescence during olfactory epithelium development in mice.


Author(s):  
Narmin Suvarli ◽  
Lukas Wenger ◽  
Christophe Serra ◽  
Iris Perner-Nochta ◽  
Jürgen Hubbuch ◽  
...  

Increasing the shelf life of enzymes and making them reusable is a prominent topic in biotechnology. The encapsulation inside hydrogel microparticles (HMPs) can enhance the enzyme’s stability by preserving its native conformation and facilitating continuous biocatalytic processes and enzyme recovery. In this study, we present a method to immobilize β-galactosidase by, first, conjugating the enzyme onto the surface of polymer nanoparticles, and then encapsulating these enzyme-conjugated nanoparticles (ENPs) inside HMPs using microfluidic device paired with UV-LEDs. Polymer nanoparticles act as anchors for enzyme molecules, potentially preventing their leaching through the hydrogel network especially during swelling. The affinity binding (through streptavidin-biotin interaction) was used as an immobilization technique of β-galactosidase on the surface of polymer nanoparticles. The hydrogel microparticles of roughly 400 μm in size (swollen state) containing unbound enzyme and ENPs were produced. The effects of encapsulation and storage in different conditions were evaluated. It was discovered that the encapsulation in acrylamide (AcAm) microparticles caused an almost complete loss of enzymatic activity. Encapsulation in poly(ethylene glycol) (PEG)-diacrylate microparticles, on the other hand, showed a residual activity of 15–25%, presumably due to a protective effect of PEG during polymerization. One of the major factors that affected the enzyme activity was presence of photoinitiator exposed to UV-irradiation. Storage studies were carried out at room temperature, in the fridge and in the freezer throughout 1, 7 and 28 days. The polymer nanoparticles showcased excellent immobilization properties and preserved the activity of the conjugated enzyme at room temperature (115% residual activity after 28 days), while a slight decrease was observed for the unbound enzyme (94% after 28 days). Similar trends were observed for encapsulated ENPs and unbound enzyme. Nevertheless, storage at −26°C resulted in an almost complete loss of enzymatic activity for all samples.


2022 ◽  
Author(s):  
Irem Avcilar-Kucukgoze ◽  
Anna Kashina

Protein arginylation, mediated by arginyltransferase ATE1, is a posttranslational modification of emerging biological importance that consists of transfer of the amino acid Arg to protein and peptide substrates. ATE1 utilizes charged tRNAArg as the donor of the arginyl group, which depends on the activity of Arg-tRNA synthetases (RARS) and is also utilized in translation. The mechanisms that regulate the functional balance between ATE1, RARS and translation are unknown. Here we addressed the functional interplay between these mechanisms using intracellular arginylation sensor in cell lines with overexpression or deletion of ATE1 and RARS isoforms. We find that arginylation levels depend on the physiological state of the cells but are not directly affected by translation activity or availability of RARS isoforms. However, displacement of RARS from the multi-synthetase complex leads to an increase in intracellular arginylation independently of RARS enzymatic activity. This effect is accompanied by ATE1 redistribution into the cytosol. Our results provide the first comprehensive analysis of the interdependence between translation, arginyl-tRNA synthesis, and arginylation.


2022 ◽  
Vol 55 (1) ◽  
Author(s):  
Fatemeh Safari ◽  
Bahman Akbari

Abstract Background Chinese hamster ovary cell line has been used routinely as a bioproduction factory of numerous biopharmaceuticals. So far, various engineering strategies have been recruited to improve the production efficiency of this cell line such as apoptosis engineering. Previously, it is reported that the caspase-7 deficiency in CHO cells reduces the cell proliferation rate. But the effect of this reduction on the CHO cell productivity remained unclear. Hence, in the study at hand the effect of caspase-7 deficiency was assessed on the cell growth, viability and protein expression. In addition, the enzymatic activity of caspase-3 was investigated in the absence of caspase-7. Results Findings showed that in the absence of caspase-7, both cell growth and cell viability were decreased. Cell cycle analysis illustrated that the CHO knockout (CHO-KO) cells experienced a cell cycle arrest in G2/M phase. This cell cycle arrest resulted in a 1.7-fold increase in the expression of luciferase in CHO-KO cells compared to parenteral cells. Furthermore, in the apoptotic situation the enzymatic activity of caspase-3 in CHO-KO cells was approximately 3 times more than CHO-K1 cells. Conclusions These findings represented that; however, caspase-7 deficiency reduces the cell proliferation rate but the resulted cell cycle arrest leads to the enhancement of recombinant protein expression. Moreover, increasing in the caspase-3 enzymatic activity compensates the absence of caspase-7 in the caspase cascade of apoptosis.


2022 ◽  
Vol 7 (1) ◽  
Author(s):  
Anqi Zheng ◽  
Lili Wu ◽  
Renyi Ma ◽  
Pu Han ◽  
Baoying Huang ◽  
...  
Keyword(s):  

2022 ◽  
pp. 42-45
Author(s):  
Юлия Юрьевна Миллер ◽  
Татьяна Федоровна Киселева ◽  
Лариса Викторовна Пермякова ◽  
Юлия Владимировна Арышева

Определяющей целью солодоращения является повышение ферментативной активности зерна. Нами предлагается способ интенсификации солодоращения пшеницы посредством применения неорганического стимулятора роста «Энерген». В исследовании использовали пшеницу Алтайской селекции трех сортов: «Алтайская 100», «Дуэт» и «Алейская». Предложенный неорганический препарат вносили при замачивании в последнюю замочную воду в количестве 0,6 г/дм и выдерживали с ним в контакте пшеницу в течение 6 ч. За данный период в ферментативной системе обработанного зерна произошли более выраженные изменения в сравнении с контрольным вариантом (необработанным зерном). К концу замачивания уровень активности ферментов опытных образцов стал выше уровня аналогичных активностей ферментов контрольных вариантов на 11,8 и 9,9 % соответственно для амилолитической и протеолитической активностей. Последующее проращивание зерна повысило ферментативную активность пшеничного солода. По окончании 7 сут данной стадии прирост амилолитической активности над активностями необработанного зерна для разных сортов составил от 31,5 до 59,0 %, протеолитической - от 97,8 до 125,4 %. При этом отмечено маловыраженное отличие показателей амилолитической и протеолитической активностей проращиваемого обработанного пшеничного солода шестых и седьмых суток ращения, что позволяет сократить продолжительность данной стадии и всего производства солода на одни сутки. Готовый пшеничный солод отличался высокой ферментативной активностью (в диапазоне для трех сортов): амилолитическая - 344,9-360,8 ед./г, протеолитическая - 324,9-257,8 ед./г, более низкой в сравнении с контрольным вариантом продолжительностью осахаривания - от 18 до 20 мин. Кроме этого, предложенный способ солодоращения позволяет использовать пшеницу с высоким содержанием белка, как, например, сорт «Алейская» с массовой долей белка 14,6 %, поскольку в процессе проращивания под стимулирующим действием неорганического препарата «Энерген» процесс протеолиза протекает более интенсивно, и в конечном солоде содержание белка снижается до 10,4 %. The defining goal of malting is to increase the enzymatic activity of grain. We propose a method for intensifying the malting of wheat through the use of an inorganic growth stimulator «Energen». The study used wheat of the Altai selection of three varieties: «Altai 100», «Duet» and «Aleyskaya». The proposed inorganic preparation was introduced during soaking into the last soak water in an amount of 0.6 g/dm and wheat was kept in contact with it for 6 hours. During this period, more pronounced changes occurred in the enzymatic system of the processed grain in comparison with the control variant (unprocessed grain). By the end of soaking, the enzyme level of the experimental samples is 11.8 and 9.9 % higher than the level of similar enzymes of the control variants, respectively, for amylolytic and proteolytic activities. The subsequent germination of grain increased the enzymatic activity of wheat malt. At the end of seven days of this stage, the increase in amylolytic activity over the activities of unprocessed grain for different varieties ranged of 31.5 to 59.0 %, proteolytic - of 97.8 to 125.4 %. At the same time, there was a little pronounced difference in the indicators of amylolytic and proteolytic activities of the germinated processed wheat malt of the sixth and seventh days of fermentation, which makes it possible to shorten the duration of this stage and the entire malt production by one day. The finished wheat malt was characterized by high enzymatic activity (in the range for three varieties): amylolytic 344.9-360.8 units /g, proteolytic 324.9-257.8 units/g, lower duration of saccharification in comparison with the control variant of 18 to 20 minutes. In addition, the proposed method of malting allows the use of wheat with a high protein content, such as the Aleyskaya variety with a mass fraction of protein of 14.6 %, since during germination under the stimulating effect of the inorganic preparation Energen, the proteolysis process proceeds more intensively, and in the final malt the protein content decreases to 10.4 %.


2022 ◽  
Vol 12 ◽  
Author(s):  
Ales Hanc ◽  
Bayu Dume ◽  
Tereza Hrebeckova

The study aims were focused on profiling eight hydrolytic enzymes by fluorescence method using a multifunctional modular reader and studying the proportion of basic microorganism groups during composting and vermicomposting of sewage sludge mixed with straw pellets in several proportions (0, 25, 50, 75, and 100%). The greatest decrease in enzymatic activity occurred in the first half of composting and vermicomposting. After 4 months of these processes, the least enzymatic activity was observed in the sludge with 50% and also 25% straw addition, indicating that straw is an important means for the rapid production of mature compost from sewage sludge. Enzymatic activity was usually less in the presence of earthworms than in the control treatment because some processes took place in the digestive tract of the earthworm. For the same reason, we observed reduced enzyme activity during fresh feedstock vermicomposting than precomposted material. The final vermicompost from fresh feedstocks exhibited less microbial biomass, and few fungi and G− bacteria compared to precomposted feedstock. The enzymatic activity during composting and vermicomposting of sewage sludge and their mixtures stabilized at the following values: β-D-glucosidase—50 μmol MUFG/h/g dw, acid phosphatase—200 μmol MUFP/h/g dw, arylsulphatase—10 μmol MUFS/h/g dw, lipase—1,000 μmol MUFY/h/g dw, chitinase—50 μmol MUFN/h/g dw, cellobiohydrolase—20 μmol MUFC/h/g dw, alanine aminopeptidase—50 μmol AMCA/h/g dw, and leucine aminopeptidase—50 μmol AMCL/h/g dw. At these and lesser values, these final products can be considered mature and stable.


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