Встановлення режимів термічної обробки ікри прісноводної риби для підвищення її мікробіологічної безпеки

Fish roe is one the most valuable food products; however it is very suitable environment for existence and reproduction of microorganisms. One of the best known methods for keeping fish roe quality is pasteurization. Currently, different pasteurization regimes are used both at temperature and time. The temperature can vary from 63 to above 100 °C. Because of the protein content in polyunsaturated fatty acids, the roe is very sensitive to temperature. Successful development of high-quality and safe food products based on freshwater fish roe is possible when it is provided with improvement and updating of heat treatment regimes. The aim of the work is to substantiate the regimes of heat treatment of freshwater fish roe on the base of its microbiological indicators. As raw material for the study, ich roe (Hypophthalmichthys) and carp roe (Cyprinus carpio) were used. In order to determine the optimum temperature of heat treatment the freshwater fish roe were subjected to heat treatment at a temperature of 60 °С, 65 °С, 70 °С, 75 °С, 80 °С for 60 minutes. The optimum temperature ensuring complete destruction of commensals and allow the permissible amount of residual microflora (MAFAnM) is determined at 70–75 °С. The duration of the heat treatment process was determined on the base of evaluation of dynamics in amount changes of MAFAnM at a temperature of 70 °С for 30, 45, 60, 75, 90 minutes. The rational duration of heat treatment is 75 minutes, which ensures the destruction of vegetative forms of microorganisms, and the amount of residual microflora is within the permissible norm.  Рибна ікра є одним з найцінніших продуктів харчування та сприятливим середовищем для існування та розмноження мікроорганізмів. Одним із найбільш відомих методів збереження якості ікри є пастеризування. На даний час використовують різні режими пастеризування як за температурою, так і за часом. Температура може коливатися від 63 °С до вище 100 °С. За рахунок вмісту білку, поліненасичених жирних кислот ікра дуже чутлива до температурного впливу. Успішна розробка якісних та безпечних харчових продуктів на основі ікри прісноводної риби можлива за умови удосконалення і уточнення режимів термічного оброблення ікри. Метою роботи є обґрунтування режимів термічного оброблення ікри прісноводної риби на основі мікробіологічних показників. В якості сировини для дослідження використовували ікру товстолобика (Hypophthalmichthys) і коропа (Cyprinus carpio). З метою визначення оптимальної температури теплового оброблення ікру прісноводної риби піддавали термічному обробленню за температури 60 °С, 65 °С, 70 °С, 75 °С, 80 °С протягом 60 хвилин. Оптимальною температурою, що забезпечувала повне знищення умовно-патогенних мікроорганізмів і допустиму кількість залишкової мікрофлори (МАФАнМ) визначено 70–75 °С. Тривалість процесу термічного оброблення встановлювали на основі оцінки динаміки зміни кількості МАФАнМ за температури 70 °С протягом 30, 45, 60, 75, 90 хвилин. Раціональною тривалість теплового оброблення є 75 хвилин, що забезпечує знищення вегетативних форм мікроорганізмів, а кількість залишкової мікрофлори знаходиться в межах допустимої норми. &nbsp

Role of Plant-Specific N-Terminal Domain of Maize CK2β1 Subunit in CK2β Functions and Holoenzyme Regulation

Protein kinase CK2 is a highly pleiotropic Ser/Thr kinase ubiquituous in eukaryotic organisms. CK2 is organized as a heterotetrameric enzyme composed of two types of subunits: the catalytic (CK2α) and the regulatory (CK2β). The CK2β subunits enhance the stability, activity and specificity of the holoenzyme, but they can also perform functions independently of the CK2 tetramer. CK2β regulatory subunits in plants differ from their animal or yeast counterparts, since they present an additional specific N-terminal extension of about 90 aminoacids that shares no homology with any previously characterized functional domain. Sequence analysis of the N-terminal domain of land plant CK2β subunit sequences reveals its arrangement through short, conserved motifs, some of them including CK2 autophosphorylation sites. By using maize CK2β1 and a deleted version (ΔNCK2β1) lacking the N-terminal domain, we have demonstrated that CK2β1 is autophosphorylated within the N-terminal domain. Moreover, the holoenzyme composed with CK2α1/ΔNCK2β1 is able to phosphorylate different substrates more efficiently than CK2α1/CK2β1 or CK2α alone. Transient overexpression of CK2β1 and ΔNCK2β1 fused to GFP in different plant systems show that the presence of N-terminal domain enhances aggregation in nuclear speckles and stabilizes the protein against proteasome degradation. Finally, bimolecular fluorescence complementation (BiFC) assays show the nuclear and cytoplasmic location of the plant CK2 holoenzyme, in contrast to the individual CK2α/β subunits mainly observed in the nucleus. All together, our results support the hypothesis that the plant-specific N-terminal domain of CK2β subunits is involved in the down-regulation of the CK2 holoenzyme activity and in the stabilization of CK2β1 protein. In summary, the whole amount of data shown in this work suggests that this domain was acquired by plants for regulatory purposes

Phosphorylation of Maize Eukaryotic Translation Initiation Factor 5A (eIF5A) by Casein Kinase 2: IDENTIFICATION OF PHOSPHORYLATED RESIDUE AND INFLUENCE ON INTRACELLULAR LOCALIZATION OF eIF5A*

Maize eukaryotic translation initiation factor 5A (ZmeIF5A) co-purifies with the catalytic α subunit of protein kinase CK2 and is phosphorylated by this enzyme. Phosphorylated ZmeIF5A was also identified after separation of maize leaf proteins by two-dimensional electrophoresis. Multiple sequence alignment of eIF5A proteins showed that in monocots, in contrast to other eukaryotes, there are two serine/threonine residues that could potentially be phosphorylated by CK2. To identify the phosphorylation site(s) of ZmeIF5A, the serine residues potentially phosphorylated by CK2 were mutated. ZmeIF5A and its mutated variants S2A and S4A were expressed in Escherichia coli and purified. Of these recombinant proteins, only ZmeIF5A-S2A was not phosphorylated by maize CK2. Also, Arabidopsis thaliana and Saccharomyces cerevisiae eIF5A-S2A mutants were not phosphorylated despite effective phosphorylation of wild-type variants. A newly developed method exploiting the specificity of thrombin cleavage was used to confirm that Ser2 in ZmeIF5A is indeed phosphorylated. To find a role of the Ser2 phosphorylation, ZmeIF5A and its variants mutated at Ser2 (S2A and S2D) were transiently expressed in maize protoplasts. The expressed fluorescence labeled proteins were visualized by confocal microscopy. Although wild-type ZmeIF5A and its S2A variant were distributed evenly between the nucleus and cytoplasm, the variant with Ser2 replaced by aspartic acid, which mimics a phosphorylated serine, was sequestered in the nucleus. These results suggests that phosphorylation of Ser2 plays a role in regulation of nucleocytoplasmic shuttling of eIF5A in plant cells