Hepatitis B virus nucleocapsid particles produced in eukaryotic cells: Properties and purification

s / Journal of Biotechnology 185S (2014) S37–S125 S71 collagen from osteoblastic-like MC3T3-E1 cells. This enzyme is a new biotechnology product (patent pending) that can be applied in several industrial, life-science and clinical research applications whereas collagen hydrolysis and tissues dissociation is required. Acknowledgements PROMETHEUS project and FEDER – COMPETE program (PTDC/AGR-CFL/113831/2009 and FCOMP-01-0124-FEDER014096) and FCT grants to AC Esteves (BPD/38008/2007) and AS DUARTE (SFRH/BPD/46290/2008). http://dx.doi.org/10.1016/j.jbiotec.2014.07.239 Hepatitis B virus nucleocapsid particles produced in eukaryotic cells: Properties and purification Karina Spunde, Alma Eihentale ∗, Anna Zajakina, Tatyana Kozlovska, Paul Pumpens Latvian Biomedical Research and Study Centre, Riga, Latvia E-mail address: kspunde@biomed.lu.lv (A. Eihentale). Hepatitis B virus core (HBc) particle is a favorite and most studied virus-like particle model for vaccine candidates generation and gene therapy tools. Nevertheless, many structural features of native HBc particles remain unclear. With an aim to study structural and functional peculiarities of native HBc particles, the stable HBc producing eukaryotic cell lines were established. To achieve the efficient HBc production, the thermoinducible noncytopathic alphavirus vector with inserted HBc gene, was generated and introduced into BHK-21 and CHO-K1 cells by electroporation and nucleofection respectively. After selection of HBc producing clones, the HBc expression was evaluated by ELISA, Western blot and fluorescentmicroscopy. HBc particle formationwas confirmed by electron microscopy. Different core particle purification protocols have been tried. Despite some achievements, moderate HBc production level inmammalian cells,was thedefining for the specificity of purification and the procedure still have to be improved. It was found, that eukaryotic HBc is phosphorylated and that the particles, contrary to the Escherichia coli produced, do not contain nucleic acid. These features undoubtedly are crucial determinants concerning core particles surface charge and stability. As a result, the stablemammalian cell linewith efficient synthesis and production of native phosphorylated HBc particles was established. http://dx.doi.org/10.1016/j.jbiotec.2014.07.240 Purification and characterization of a cold alkaline protease from a psychrophilic Pseudomonas aeruginosa HY1215 Hao Jianhua ∗, Sun Mi Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, China E-mail address: haojh@ysfri.ac.cn (H. Jianhua). A novel alkaline protease was purified from Pseudomonas aeruginosa HY1215 using ammonium sulphate, DEAE-Sepharose andSephacryl S-200gel filtration chromatographic techniques. The protease had a relative molecular weight of 32.8 kDa by SDS-PAGE, and the optimal temperature and pH for excellent stability and activity were determined as 25 ◦C and 10.0, respectively. Within the pH range of 7.0–11.0, the protease had a good stability, which could retain more than 80% of its original activity; in the temperature range of 15–35 ◦C, the protease had a higher activity, and its activity at 20 ◦C could amount to 85% of the maximum activity at 25 ◦C. Besides, the enzyme activity showed a valuable stability towards several commercially available surfactants (Tween-80, Tween-40 and Triton X-100) and bleaches (H2O2) even when their concentrations ranged up to 2.0% and 1.6%. http://dx.doi.org/10.1016/j.jbiotec.2014.07.241 Pleurotus ostreatus laccases: New media for maximum production at minimum cost Lucia Guarino1,∗, Vincenzo Lettera2, Giovanni Sannia1 1 Department of Chemical Science, University of Naples, Federico II, Naples, Italy 2 Biopox s.r.l., via Salita Arenella, 9, Naples, Italy E-mail address: lucia.guarino@unina.it (L. Guarino). Enzymesarebeing rigorously and systematically developed, as economically viable and environment-friendly industrial biocatalysts. As a fact the global market for industrial enzymes was estimated $3.9 billion in 2011. BCC projects this market to grow at a compounded annual growth rate (CAGR) of 9.1% to reach $6 billion by 2016. Pleurotus ostreatus, an edible basidiomycete, produces a wide range of extracellular industrially relevant enzymes such as laccases. Themost significant limit of enzyme commercialization is the high production cost that is highly influenced by culture broth. In order to optimize laccases production (costs and productivity) by P. ostreatus two approaches were used. Trough a one factor at time approach rapeseed cake (a waste product coming from rapeseed oil extraction processes), a lignosulphonates mixture, copper sulphate, and yeast extract were selected as factors that affect the productivity and the costs. The interactions between the different factors composing growth medium were evaluated trough a statistical multi-factorial approach such as the Response Surface Methodology (RSM). The two approaches lead to a laccase production level close to 180,000U/L at a cost of about 8×10−7 D/U. http://dx.doi.org/10.1016/j.jbiotec.2014.07.242 Food & Feed Biotechnology Quality of cow milk and for some cheeses assortments Ratu Nicoleta Roxana ∗, Usturoi Giorgi Marius, Avarvarei Vlad Bogdan Department of Management of Animal Productions, University of Agricultural Sciences and Veterinary Medicine “Ion Ionescu de la Brad”, Iasi, Romania E-mail address: roxana.ratu@gmail.com (R.N. Roxana). In the current paper the quality of cheeses manufactured in a specialized unit for milk processing is analyzed. Naturally for a better appreciation of their quality has been imposed and analysis of raw milk in terms of its sensorial and physical-chemical properties. Thus from measurements made on raw milk was obtained an average density of 1.027±0.010g/cm3 while the fat content had a mean of 3.693±0.076% and a variation coefficient of 5.501% which indicates a homogeneous character. The tests were made for determining acidity and protein, the average for protein content being2.955±0.026. The analyzed assortments of cheeseswere representedbyCheese “Rucăr” and “SmokedCheese” for bothbeing analysed acidity, salt, water, fat and protein content. For the first analyzed assortment we obtain an average value for acidity of