• Journal of Microbiology and Biotechnology
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• v.17 no.4
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• pp.579-585
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• 2007

Formation of inclusion bodies is usually observed when foreign proteins are overexpressed in E. coli. The formation of inclusion bodies might be prevented by lowering the rate of protein synthesis, and appropriate regulation of the protein expression rate may lead to the soluble expression. In this study, human growth hormone (rhGH) was expressed in a soluble form by slowing down the protein synthesis rate, which was controlled in the transcriptional and translational levels. The transcriptional level was controlled by the regulation of the amount of RNA polymerase specific to the promoter in front of the rhGH gene. For lowering the rate of translation, the T7 transcription terminator-deleted vector was used to synthesize the longer mRNA of the target gene because the longer mRNA is expected to reduce the availability of tree ribosomes. In both methods, the percentage of soluble expression increased when the expression rate slowed down, and more than 93% of rhGH expressed was a soluble form in the T7 transcription terminator-deleted expression system.

• Journal of the Korean Society of Food Science and Nutrition
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• v.2 no.1
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• pp.1-21
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• 1973

The muscle of abalone, Notohaliotis discus (REEVE), and top-shell, Turbo cornutus Solander, were examined for protein composition. Then paramyosins which are known as one of the important structural protein of the muscle fibrils were isolated from the both muscle and their physico-chemical properties such as solubility, salting-out behaviour, intrinsic viscosity, ATPase activity, etc. involving amino acid composition and N-terminal amino acid residues were investigated to elucidate phylogenie characteristics more intensively from the viewpoint of comparative biochemistry. The analysis of protein composition resulted in the following estimations: abalone muscle; water-soluble protein of 22 %, salt-soluble protein, 34%, alkali-soluble protein, 20%, and stroma protein, 24%, and top-shell muscle; water-soluble protein of 16%, salt-soluble protein, 30%, alkali-soluble protein, 29%, and stroma protein, 25%, respectively. It is demonstrated in sedimentation analysis that paramyosin and myosin-actomyosin account for approximately 65% and 35% of the salt-soluble protein of abalone, and that the composition of both sediments in top-shell was approximately 70% and 30%, respectively. The ultracentrifugally homogenous paramyosins isolated essentially according to Bailey's ethanol-dried method from both of the muscle showed a $S^{\circ}_{20,w}$ of 3. 14s for abalone and a $S^{\circ}_{20,w}$ of 3.50s for top-shell. The both paramyosins were commonly rich in arginine, aspartic acid, and glutamic acid, while scarcely contained proline and tryptophan, in rough accord with the other paramyosins thus far reported. It is clear that these gastropod paramyosins showed of having the characteristic N-terminal amino acid residues such as N-aspartic acid, N-valine, N-serine, and N-threonine in common. The abalone paramyosin completely salted in with KCl beyond $0.35{\mu}$ and the top-shell paramyosin beyond $0.30{\mu}$. The abalone paramyosin was salted-out between 18% and 30% saturation of ammonium sulphate and the top-shell paramyosin between 22% and 29% saturation. The intrinsic viscosities at abalone and top-shell paramyosins at $25^{\circ}C$ were estimated respectively to be 3.1 dl/g and 2.6 dl/g showing somewhat higher than the values for some other paramyosins from lamellibranchs. In regard with the ATPase activity, the para myosin specimens did not exhibit any significant activity over through the pH conditions of 5 to 9.5. irrespective of the presence of $Ca^{++}$ or $Mg^{++}$. So was the case with the abalone paramyosin prepared by a slightly modified Bailey's wet-extraction method.

• International Journal of Industrial Entomology and Biomaterials
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• v.7 no.1
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• pp.45-49
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• 2003

Foliar treatment with different concentrations of potassium chloride (KCl) to mulberry plants resulted in higher level of total chlorophyll, total sugars, soluble protein, in vivo nitrate reductase activity (NRA), net photosynthetic rate (NPR), pWUE and leaf yield. Optimal concentration was found to be 10.0 mM KCl with limited irrigation provided in the mulberry plantation planted in 90 ${times}$ 90 cm spacing. The deleterious effect of soil moisture stress condition has been found to be overcome by KCl foliar spray twice at 15 days interval. Regression and correlation coefficients were analyzed, and a strong positive correlation was found between chlorophyll and total sugars, soluble protein and in vivo nitrate reductase activity, leaf dry weight and net photosynthetic rate and pWUE and net photosynthetic rate.

• KOREAN JOURNAL OF CROP SCIENCE
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• v.23 no.1
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• pp.1-4
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• 1978

The change of soluble protein components in leaf discoloration of rice plant was investigated in the Growth Cabinet with various temperature conditions. The ratio between high molecular soluble protein and low molecular soluble protein was high under high temperature condition, while low under low temperature condition, and also lower in Indica type varieties than Japonica type variety.

• Microbiology and Biotechnology Letters
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• v.18 no.3
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• pp.239-243
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• 1990

In this study the authors investigated the distribution of cadmium accumulated in cadmium-iun tolerant Hansenula anomala B-7 cells and also the effect of cadmium on protein synthesis. 84.9% of the cadmium accumulated was distributed in the soluble fraction (cytosol, etc.). The intracellular protein content was decreased by cadmium (1,000 $\mu g$/ml), but the content of soluble protein preeipitated by ammonium sulfate (30-75% saturation) was increased compared with the content of it obtained from the cells grown without cadmium. Furthermore, in the cells grown with 1,000 $\mu g$/ml of cadmium t h higher molecular weight soluble protein was increased compared with the cells grown without caa, mium, but the lower molecular weight soluble protein was decreased. These results suggested that the protein synthesis was inhibited by cadmium, but synthesis of higher molecular weight soluble protein was remarkably stimulated by cadmium.

• Biotechnology and Bioprocess Engineering:BBE
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• v.8 no.2
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• pp.89-93
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• 2003

human renin binding protein (hRnBp), showing N-acetylglucosamine-2-epimerase activity, was over-expressed in E. coli, but was mainly present as an inclusion body. To improve its solubility and activity, ubiquitin (Ub), thioredoxin (Trx), maltose binding protein (MBP) and NusA, were used as fusion partners. The comparative solubilities of the fusion proteins were, from most to least soluble: NusA, MBP, Trx, Ub. Only the MBP fusion did not significantly reduce the activity of hRnBp, but enhanced the stability. The Origami (DE3), permitting a more oxidative environment for the cytoplasm in E. coli; helped to increase its functional activity.

• International Journal of Industrial Entomology and Biomaterials
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• v.8 no.1
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• pp.67-71
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• 2004

Ten improved mulberry varieties (Vl, C1730, C2016, C2017, Anantha, RFS-175, Thallaghatapura, Vishala, S1 and S1635) were evaluated through enzyme assay and estimation of soluble protein content followed by regression analysis, grown under irrigated conditions in the alluvial soils of Gangetic plains of West Bengal in India for five successive crops in a year, The nitrate reductase (EC No. 1.6.6.1) activity (NRA, $\mu$mol N $O_2$- $h^{-1}$ $g^{-1}$ fr, wt.), total soluble protein (mg $g^{-1}$ fr, wt.) was estimated which showed to vary significantly in the tested varieties. In addition to these, the other parameters like unit leaf fresh and dry weight (g), moisture ％, unit leaf area ($\textrm{cm}^2$), specific leaf weight (g c $m^{-2}$ ), total soluble sugar (mg $g^{-1}$ fr, wt.), leaf yield/plant (kg), shoot yield/plant (kg) and net photosynthetic rate (NPR, $\mu$㏖ $m^{2}$ $s^{-1}$ ) were also studied which showed to vary significantly in tested varieties. Among them, S1635, haying higher NRA (13.25 $\mu$㏖ N $O_2$- $h^{-l}$ $g^{-1}$ fr, wt.), total soluble protein (39.63mg $g^{-1}$ fr, wt.), NPR(16.66 $\mu$㏖ $m^{-2}$ $s^{-1}$ ), total soluble sugar (48.44 mg $g^{-1}$ fr. wt.), leaf yield/plant (0.689 kg) and shoot yield/plant (1.135 kg) showed its superiority over other tested varieties. Regression and correlation coefficients were analysed, and a strong positive correlation was found between NRA ＆ total soluble protein, NRA ＆ NPR, NRA ＆ total soluble sugar, NRA af unit leaf weight, NRA ＆ specific leaf weight, NRA ＆ leaf yield/plant, NRA ＆ shoot yield/plant, NPR ＆ leaf yield and NPR ＆ specific leaf weight.t.

• Korean Journal of Food Science and Technology
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• v.30 no.6
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• pp.1388-1393
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• 1998

Traditional Andong sikhe was produced by fermenting L. bulgaricus LBS 47 and S. cerevisiae SCS 5. The changes of nitrogen compound and amino acid during fermentation and storage were investigated. Crude protein was increased until 4days, the main fermentation period. Amino form nitrogen increased up to 37.50 mg% at the 2nd day of fermentation and the product tasted best at this time. Water soluble and salt soluble protein decreased during fermentation. Proline and aspartic acid were the two major free amino acids. The free methionine increased while the free lysine decreased in the process of fermentation. The amino acids of water soluble protein and salt soluble protin were totally 17 kinds. The major amino acids of water soluble and salt soluble protein were glutamic acid and aspartic acid. The arginine content of salt soluble protein increased as the fermentation proceeded.

• Asian-Australasian Journal of Animal Sciences
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• v.16 no.10
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• pp.1460-1468
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• 2003

An experiment was conducted to quantify the flow of soluble non-ammonia nitrogen (SNAN) in the liquid phase of ruminal (RD) and omasal digesta (OD), and to investigate diurnal pattern in SNAN flow in OD. Five ruminally cannulated Finnish-Ayrshire dairy cows in a $5{\times}5$ Latin square design consumed a basal diet of grass silage and barley grain, and that supplemented with four protein feeds (kg/d DM basis) as follows: skimmed milk powder (2.1), wet distiller' solubles (3.0), untreated rapeseed meal (2.1) and treated rapeseed meal (2.1). Ruminal digesta was sampled using a vacuum pump, whereas OD was collected using an omasal sampling system at 1.0 h interval during a 12 h feeding cycle. Both RD and OD were acidified, centrifuged to remove microbes and precipitated with trichloroacetic acid followed by centrifugation. The SNAN fractions (free amino acid (AA), peptide and soluble protein) in RD and OD were assessed using ninhydrin assay. Free AA, peptide and soluble protein averaged 60.0, 89.4 and 2.1 g/d, respectively, for RD, and 81.8, 121.5 and 2.5 g/d, respectively, for OD. Although free AA flow was relatively high, mean peptide flow was quantitatively the most important fraction of SNAN, indicating that degradation of peptide to AA rather than hydrolysis of soluble protein to peptide or deamination may be the most limiting step in rumen proteolysis. Diurnal pattern in flow of peptide including free AA in OD during a 12 h feeding cycle peaked 1 h post-feeding, decreased by 3 h post-feeding and was relatively constant thereafter. Protein supplementation showed higher flow of peptide including free AA immediately after feeding compared with no supplemented diet. There were no differences among protein supplements in diurnal pattern in flow of peptide including free AA in OD.

• Microbiology and Biotechnology Letters
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• v.49 no.2
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• pp.148-156
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• 2021

Single-chain antibodies against epidermal growth factor receptor variant III (EGFRvIII) are potentially promising agents for developing antibody-based cancer treatment strategies. We described in our previous study the successful expression of an anti-EGFRvIII scFv antibody in Escherichia coli. However, we could also observe the formation of insoluble aggregates in the periplasmic space, limiting the production yield of the active product. In the present study, we investigated the mechanisms by which growth conditions could affect the expression of the soluble anti-EGFRvIII scFv antibody in small-scale E. coli NiCo21(DE3) cultures, attempting to maximize production. The secreted scFv molecules were purified using Ni-NTA magnetic beads and protein characterization was performed using SDS-PAGE and western blot analyses. We used the ImageJ software for protein quantification and determined the antigen-binding activity of the scFv antibody against the EGFRvIII protein. Our results showed that the highest percentage of soluble scFv expression could be achieved under culture conditions that combined low IPTG concentration (0.1 mM), low growth temperature (18℃), and large culture dish surface area. We found moderate-yield soluble scFv production in the culture medium after lactose-mediated induction, which was also beneficial for downstream protein processing. These findings were confirmed by conducting western blot analysis, indicating that the soluble, approximately 30-kDa scFv molecule was localized in the periplasm and the extracellular space. Moreover, the antigen-binding assay confirmed the scFv affinity against the EGFRvIII antigen. In conclusion, our study reveals that low-speed protein expression is preferable to obtain more soluble anti-EGFRvIII scFv protein in an E. coli expression system.