• Journal of Microbiology and Biotechnology
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• v.19 no.7
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• pp.727-733
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• 2009

Heat shock protein 70 kDa (hsp70), a molecular chaperone involved in folding of nascent proteins, has been studied for its ability to activate innate and specific immunity. High purity hsp70 preparation is generally required for immunization experiments, because endotoxins and other immunologically active contaminants may affect immune responses independently of hsp70. We have developed a novel modification of E. coli-expression medium that enabled a simple two-step production and purification method for endotoxin-free recombinant hsp70. During Ni-NTA-based affinity purification of hsp70, a contaminating protein from host E. coli cells, L-glutamine-n-fructose-6-phosphate aminotransferase (GFAT), was identified. By testing various compounds, supplementation of growth medium with a GFAT metabolite,N-acetylglucosamine, was found to reduce GFAT expression and increase the total hsp70 yield five times. The new protocol is based on column purification of His-tagged hsp70 protein produced by E. coli with the modified medium, followed by endotoxin removal by Triton X-114 extraction. This approach yielded hsp70 with high purity and minimal endotoxin contamination, making the final product acceptable for immunization experiments. In summary, a simple modification of growth medium allowed production of recombinant mouse hsp70 in high yield and purity, thus compatible with immunological studies. This protocol may be useful for production of other Histagged proteins expressed in E. coli.

• 한국신재생에너지학회:학술대회논문집
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• 2007.06a
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• pp.69-72
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• 2007

The objective of this work was to study the properties of purification of two liquid phase for exclusion of impurities in each phase. The experiments for process variables were carried out in the temperature range($H_{2}SO_{4}$ phase: $413{\sim}513$ K, $HI_{x}$ phase: $353{\sim}453$ K) and in the $N_{2}$ flow rate range($H_{2}SO_{4}$, $HI_{x}$ phase: $50{\sim}200$ mL/min). As the results, it is appeared that the principles of $H_{2}SO_{4}$ phase purification was due to stripping, evaporation and reverse bunsen reaction and $HI_{x}$ phase purification was due to stripping and reverse bunsen reaction. In purification of $H_{2}SO_{4}$ phase, the concentration rate of $H_{2}SO_{4}$ phase was controled by temperature but the temperature had few effects on yield of $H_{2}SO_{4}$. In purification of $HI_{x}$ phase, we observed products of side reactions($H_{2}S$, S) over 433 K. The purity of $HI_{x}$ phase was increased with increasing $N_{2}$ flow rate because impurites were decreased with increasing conversion of reverse reaction.

• Journal of Environmental Science International
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• v.33 no.2
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• pp.187-191
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• 2024

This study aimed to investigate the effect of treating environmental purification insect larvae to pig manure on crude ash contents and ammonia production. The experiment set up consisted go two groups: 1 kg of each 3^rd instar TM (Tenebrio molitor) and 3^rd instar PBS (Protaetia brevitarsis seulensis) larvae in Experiment 1 or 3^rd and 4^th instar of HI (Hermetia illucens L.) larvae in Experiment 2 were treated with 5 kg of pig manure. In Experiment 1, the crude ash content was higher in TM larvae-treated pig manure at days 0 and 5 (p>0.05), but was similar to that in PBS larvae-treated pig manure over (p>0.05). Ammonia production was observed at day 0 of TM and PBS larvae-treated pig manure (p<0.05), but did not occur thereafter. For Experiment 2, there was significant difference in crude ash content of 3^th and 4^th instar HI larvae-treated pig manure on day 15. Additionally, ammonia production was found in 3^th and 4^th instar HI larvae-treated pig manure at days 0 and 5, but did not continue over time. In conclusion, treating TM, PBS and HI to pig manure changed the crude ash contents and reduced ammonia through the ability to decompose pig manure. Thus, environmental impact can be minimized using environmental purification insect larvae.

• BMB Reports
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• v.37 no.4
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• pp.387-393
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• 2004

The L-asparaginase (E. C. 3. 5. 1. 1) enzyme was purified to homogeneity from Pseudomonas aeruginosa 50071 cells that were grown on solid-state fermentation. Different purification steps (including ammonium sulfate fractionation followed by separation on Sephadex G-100 gel filtration and CM-Sephadex C50) were applied to the crude culture filtrate to obtain a pure enzyme preparation. The enzyme was purified 106-fold and showed a final specific activity of 1900 IU/mg with a 43% yield. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of the purified enzyme revealed it was one peptide chain with $M_r$ of 160 kDa. A Lineweaver-Burk analysis showed a $K_m$ value of 0.147 mM and $V_{max}$ of 35.7 IU. The enzyme showed maximum activity at pH 9 when incubated at $37^{\circ}C$ for 30 min. The amino acid composition of the purified enzyme was also determined.

• Journal of Microbiology and Biotechnology
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• v.11 no.2
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• pp.204-210
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• 2001

Paclitaxel can be produced in high yield and with a high degree of purify from plant cell cultures of Taxus chinensis. The complete purification method was systematically established and described. This method was an efficient procedure for the purification of paclitaxel from crude paclitaxel, consisting or reverse-phase chromatography, followed by a normal-phase chromatography. The two-stage HPLC purification scheme serves as an effective and economical approach for resolving paclitaxel from complex mixtures of taxoids, with high purify (>99%) and low impurities (<0.1%). The process is readily scalable to a pilot plant and eventually to a production environment where multikilogram quantities of material are expected to be produced. The process has been optimized to minimize solvent usage, complexity, and operating costs.

• Korean Journal of Plant Tissue Culture
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• v.27 no.4
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• pp.283-297
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• 2000

Seeds are an ideal repository for recombinant proteins in molecular farming applications. However, in order to use plant seeds efficiently for the production of such proteins, it is necessary to understand a number of fundamental biological properties of seeds. This includes a full understanding of promoters which function in a seed-specific manner, the subcellular targeting of the desired polypeptide and the final form in which a protein is stored. Once a biologically active protein has been deposited in a seed, it is also critical that the protein can be extracted and purified efficiently. In this review, these issues are examined critically to provide a number of approaches which may be adopted for production of recombinant proteins in plants. Particular attention is paid to the relationship between subcellular localization and protein extraction and purification. The robustness and flexibility of seed-based production is illustrated by examples close to or already in commercial production.

• Membrane Journal
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• v.34 no.2
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• pp.124-131
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• 2024

Viruses have various applications in the biopharmaceutical industry. They are used in pesticide production, production of vaccines, gene transfers, cancer therapeutics, and more. The downstream processing of viruses is an essential step for their biological and pharmaceutical applications. Among the various processes, the purification of viruses is critical. Membrane chromatography plays a vital role in this process. While ion exchange membrane chromatography is a primarily used method, it has various limitations regarding size exclusion and insufficient purification. Also, it cannot be applied to the rapidly changing strains of viruses such as influenza. This review examines various improved methods of membrane chromatography or alternatives. It focuses on purification, viral recovery rates, and scalability of the methods.

• Korean Journal of Microbiology
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• v.37 no.1
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• pp.21-27
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• 2001

Recently many investigators have devoted considerable attention to the production and purification of PA for antigens, and the preparation of new synthetic medium (RM medium) have solved to increase the yields of the PA but, the low sensitivity of the PA to detect B. anthracis infections has remained as a problem to be solved. This study was undertaken to evaluate the yields of the PA from culture filtrates of B. anthracis Sterne $34F_2$ strain in modified RM medium in which 10 g/l of $NaHCO_3$ and 10g/l of glucose were replaced by 8 g/l and 5 g/l, and the first purification step of PA from culture broth was used hydroxyapatite. The PA was purified by hydroxyapatite column chromatography, DEAE-Sepharose CL-4B column chromatography and Toyo-pearl gel filtration chromatography. The yield of PA from the modified RM medium, 8.6 mg/l.

• 한국가스학회:학술대회논문집
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• 2005.10a
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• pp.141-145
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• 2005

DME (Di-Methyl Ether) is a new clean fuel and an environmental-friendly energy resource, also is recently increasing with an alternative interest because of the industrial use. DME has been shown to have excellent properties as a diesel fuel giving emission level better than ULEV standard. So it has been attracting considerable as an alternative diesel fuel. In this study, we carried out simulation of separation and purification process of demo plant for 101on per day DME production, which cause the effect that is important in productivity, from operation results of pilot plant for 50kg per day DME production. The liquefied stream, which was separated by gas-liquid separator after DME reactor, includes $CO_2$, DME, Methanol and $H_2O$. We established three distillation columns for separation and purification of the stream. $CO_2$ was extracted from the stream by first distillation column, DME was extracted by second column and Methanol was extracted by third column. We investigated and analyzed the effect which the actual operation variables cause in efficiency of process and optimized process, finally we got the DME of purity $100\%$.

• Microbiology and Biotechnology Letters
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• v.13 no.1
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• pp.65-70
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• 1985

The production media, partial purification and the enzymatic characteristics of laccase from Pleurotus ostreatus were investigated. Among various media tried the double strength onion media showed much higher production of laccase compared to glucose and/or CMC added media. The laccase from Pleurotus ostreatus has the optimum pH of 5.94 for the activity and appears to be stable at relatively broad pH spectrum (4.7-8.7). The experiments on the temperature stability shows that above 90% activity could be preserved after holding at $30^{\circ}C$ for 40 min., 60% activity was remained after 40 min. at $50^{\circ}C$. The Km value of the laccase from Pleurotus ostreatus was estimated to be 3.209mM and the molecular weight of laccase was estimated to be 55,000 by SDS-Polyacrylamide gel electrophoresis.