• Journal of Life Science
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• v.26 no.6
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• pp.698-704
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• 2016

The intracellular transport of organelles and protein complexes is mediated by kinesin superfamily proteins (KIFs). The first kinesin, kinesin 1, was identified as a molecular motor protein that moves various organelles and protein complexes along the microtubule rails in cells. Kinesin 1 is a tetramer of two heavy chains (KHCs, also called KIF5s) and two kinesin light chains (KLCs). KIF5s interact with many different proteins through their tail region, but their binding proteins have not yet been fully identified. To identify the interaction proteins for KIF5A, we performed yeast two-hybrid screening and found a specific interaction with ferritin heavy chain (Frt-h), which has a role in iron storage and detoxification. Frt-h bound to the amino acid residues between 800 and 940 of KIF5A and to other KIF5s in the yeast two-hybrid assay. The coiled-coil domain of Frt-h is essential for interaction with KIF5A. In addition, ferritin light chain (Frt-l) interacted with KIF5s in the yeast two-hybrid assay. In addition, these proteins showed specific interactions in the glutathione S-transferase (GST) pull-down assay. An antibody to KHC specifically co-immunoprecipitated Frt-h and Frt-l from mouse brain extracts. These results suggest the kinesin 1 motor protein may transport the ferritin complex in cells.

• Journal of Life Science
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• v.29 no.3
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• pp.369-375
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• 2019

Microtubules form through the polymerization of ${\alpha}-$ and ${\beta}-tubulin$, and tubulin transport plays an important role in defining the rate of microtubule growth inside cellular appendages, such as the cilia and flagella. Heterotrimeric kinesin 2 is a molecular motor member of the kinesin superfamily (KIF) that moves along the microtubules to transport multiple cargoes. It consists of two motor subunits (KIF3A and KIF3B) and a kinesin-associated protein 3 (KAP3), forming a heterotrimeric complex. Heterotrimeric kinesin 2 interacts with many different binding proteins through the cargo-binding domains of the KIF3s, but these binding proteins have not yet been specified. To identify these proteins for KIF3A, we performed yeast two-hybrid (Y2H) screening and found a specific interaction with ${\beta}2-tubulin$ (Tubb2), a microtubule component. Tubb2 was found to bind to the cargo-binding domain of KIF3A but did not interact with KIF3B, KIF5B, or kinesin light chain 1 in the Y2H assay. The carboxyl-terminal region of Tubb2 is essential for interaction with KIF3A. Other Tubb isoforms, including Tubb1, Tubb3, Tubb4, and Tubb5, also interacted with KIF3A in the Y2H screening. However, ${\alpha}1-tubulin$ (Tuba1) did not interact with KIF3A. In addition, an antibody to KIF3A specifically co-immunoprecipitated the KIF3B and KAP3 associated with Tubb2 from mouse brain extracts. In combination, these results suggest that a heterotrimeric kinesin 2 motor protein is capable of binding to tubulin and may transport it in cells.

• Journal of Life Science
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• v.18 no.8
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• pp.1059-1065
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• 2008

KIF21A is a member of the Kinesin superfamily proteins (KIFs), which are microtubule-dependent molecular motors, anterograde axonal transporters of cargoes. Recently, congenital fibrosis of the extraocular muscles 1 (CFEOM1) has been shown to result from a small number of recurrent heterozygous missense mutations of KIF21A. CFEOM1 results from the inability of mutated KIF21A to successfully deliver cargoes to the development of the occulo-motor neuron or neuromuscular junction. Here, we used an yeast two-hybrid system to identify a protein that interacts with the WD-40 repeat domain of KIF21A and found a specific interaction with Purkinje cell protein-2 (Pcp-2), a small protein also known as L7. Pcp-2 protein bound to the WD-40 domain of KIF21A and KIF21B but not to other KIFs in yeast two-hybrid assays. In addition, this specific interaction was also observed in the glutathione S-transferase pull-down assay. An antibody to Pcp-2 specifically co-immunoprecipitated KIF21A associated with Pcp-2 from mouse brain extracts. These results suggest that Pcp-2 may be involved in the KIF21A-mediated transport as a KIF21A adaptor protein.

• KSBB Journal
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• v.22 no.1
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• pp.16-21
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• 2007

Although Energy demands of modern society increase rapidly, current energy would be exhausted shortly. Therefore development of bio-ethanol production process from cellulose containing materials was extremly demanded. Therefore development of highly functional cellulase is requisite for this purpose. In this study cellobio-hydrolase (CBH1) gene from Trichorderma reesei was used to increase cellulase activity by directed evolution and highly functional cellobio-hydrolase was obtained and characterized.

• Journal of the Korean Society of Food Science and Nutrition
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• v.37 no.10
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• pp.1363-1368
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• 2008

The extraction and quantitative analysis conditions for astaxanthin from Haematococcus pluvialis, and the structural characteristics of H. pluvialis extract, H. pluvialis hydrolysate and synthetic astaxanthin were investigated using UV/visible and FT-IR spectrometers. Astaxanthin was dissolved in methanol, and then treated to enhance the solubility by sonication for 45 min. With sonication pretreatment, the solubility of astaxanthin increased up to 1.5 times compared to that without sonication. The extracts were hydrolyzed by cholesterol esterase for the analysis of H. pluvialis extract containing astaxanthin ester. A HPLC method using reverse phase C18 column with methanol-water (95:5, v/v) as mobile phase was developed to analyze astaxanthin. After hydrolysis, the absorption spectrum of H. pluvialis hydrolysat was changed to similar pattern to synthetic astaxanthin, confirming the extraction and analysis condition of astaxanthin from H. pluvialis.

• Korean Journal of Food Science and Technology
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• v.25 no.6
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• pp.689-693
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• 1993

${\alpha}-galactosidase$ releases galactoside from raffinose and stachyose which are the major sugars in soybean, Although raffinose and stachyose were known as flatulence factors, these sugars were recently claimed as bifidus factors. In this experiment we studied the properties of ${\alpha}-galactosidase$ and its production from Bifidobacterium sp. Int-57. Int-57 produced higher level of ${\alpha}-galactosidase$ than other intestinal bacteria. The production of ${\alpha}-galactosidase$ was greater when grown on raffinose compared with other carbohydrates tested. Partially purified ${\alpha}-galactosidase$ was obtained after sonication of harvested cell pellet followed by DEAE-cellulose chromatography and Sepharose CL-6B gel filtration, and assayed using PNP-${\alpha}-galactosidase$ as a substrate. Optimum pH for activity was 7.0 and optimum temperature was $40^{\circ}C$. At 5 mM concentration of metal ions, $CoCl_{2}\;and\;CuCl_{2}$ and inhibited the enzyme activity by 33% and 21% respectively. The enzyme was shown to hydrolyse genuine substrates, i.e. raffinose and stachyose.

• Journal of the Korean Chemical Society
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• v.39 no.6
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• pp.459-465
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• 1995

Acetolactate Synthase (ALS) was partially purified from the yeast and its basic biochemical studies were carried out. Yeast was grown in the minimum media containing 0.5% glucose, 51 mM $K_2HPO_4$, 22 mM $KH_2PO_4$, 8 mM $(NH_4)2SO_4,\;0.4\;m M\;MgSO_4$ for 18 hours at 37 $^{\circ}C$. The cell was ruptured in the buffer (20 mM phosphate buffer pH 7.0, 0.1 mM TPP, 0.5 mM DTT, 1 ${\mu}M$ FAD, and 1 mM MgCl_2$) following an overnight suspension. The supernatant fraction was collected from $10,000{\times}g$ and the enzyme was further purified by ammonium sulfate fractionation, DEAE-Sephacel chromatography and leucine-agarose chromatography. The enzyme activity was measured under the various conditions by the function of protein concentration, time, temperature, pH, and substrate. The optimum temperature was found to be 50$^{\circ}C$, optimum pH 8.0∼8.5. The kinetic parameters, $K_m\;and\;V_{max}$ were 8.4 mM and 17.9 nmol/mg/min respectively. Stability of the enzyme was studied with ethylene glycol and glycerol added to the enzyme solution. Both ethylene glycol and glycerol improved the enzyme stability up to 50%. The study of feedback inhibition showed that valine was a strong inhibitor while leucine was a weak inhibitor.

• Journal of the Korea Organic Resources Recycling Association
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• v.29 no.4
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• pp.41-46
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• 2021

In order to investigate the degree of sewage sludge solubilizaion using ultrasonic(US) combined with calcium peroxide(CaO₂), VSS reduction rate, solubilization rate, SCOD/VSS ratio, SCOD increasing rate, LB-EPS(Loosely-Bound EPS) and TB-EPS(Tightly-bound EPS) were measured. US was compared as a control. Solubilization rate increased by 23.4% under US and increased by 50.7% under US/CaO₂(0.05 g CaO₂/g VSS). and also, at the same conditions, VSS reduction rate increased by 7.1% and 17.7%, respectively. SCOD increasing rate from 10 to 90 minutes was 0.0151 min^-1 under US/CaO₂(0.02 g CaO₂/g VSS). TB-EPS decreased by 36.4% under US and decreased by 59.0% under US/CaO₂(0.05 g CaO₂/g VSS). TB-EPS decreased during first 10 minutes and then decreased slowly until 90 minutes. There was no significant difference in TB-EPS decrease according to the dosage of calcium peroxide.

• Horticultural Science & Technology
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• v.30 no.6
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• pp.700-708
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• 2012

In order to optimize a method to determine the firmness of Chinese cabbage, hardness of midrib tissues was examined based on their chronological order of emergence. Texture measurement using volodkevich bite jaws gave a consistent and highest regression ($r^2=0.85$) between firmness and the order of leaf emergence, while blade set, cylinder probe, and crisp fracture support rig showed a lower coefficient of determination. Thickness of midrib tissue within an individual head from 16 cultivars of Chinese cabbage was positively correlated with the order of emergence, becoming thinner toward inner leaves. Mean thickness of midrib tissue from the head ranged from 7.74 mm for 'CR-shingshing' and 9.28 mm for 'Norangyeorum'. The covariance of leaf thickness within a head was highly cultivar-dependent, ranging from 23.6% for 'Chihili' and 5.8% for 'Bulam'. Firmness of the midrib tissue, defined as maximum peak height per tissue thickness, became higher from outer to inner leaves, showing $2^{nd}$ order of regression. Mean firmness of the midrib tissue from individual head varied from 1.58 N for 'Rangno' to 3.46 N for 'CR-shingshing'. The $10^{th}$ or $11^{th}$ leaf brought the best correlation coefficient (r = 0.81) between firmness of an individual leaf and the mean firmness of the entire leaves in a head, suggesting a reliable and rapid method to estimate the firmness of a head in lieu of examining all leaves in the head. The relationship between firmness of midrib tissue and dry mass ($r=0.70^{**}$) as well as cell wall content ($r=0.58^*$) of the head were positively correlated. Results obtained from the present study suggested that a new method to determine midrib firmness would enable to clarify the relationship between textural quality of fresh Chinese cabbage and their processed product, 'Kimchi'. It will also be important to apply this method to screen textural quality of various genotypes under breeding programs.

• Applied Biological Chemistry
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• v.26 no.4
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• pp.222-230
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• 1983

This study was designed to develop a process for the immobilization of xylose isomerase(D-xylose ketol isomerase, EC 5.3.1.5) from Streptomyces griseolus previously isolated by the authors and its application on a pilot plant scale for the production of high fructose corn syrup. The biomass which has endo-excreted xylose isomerase was homogenized under a pressure of $500kg/cm^2$ and 90.8% of the enzyme recovery of the native activity was obtained as compared to 54.7% recovery by the lysozyme treatment. Ionic bonding method was adopted for the enzyme immobilization due to its many reported merits. It was found that the porous resins such as Diaion HP 20, Duolite A-7, Amberlite IRA 93 and 94 were effective in immobilizing the enzyme. In addition, it was disclosed that the regeneration form of $BO_4--$ is effective for Amberlite IRA 93 and $HCO_3-$ for Diaion HP 20. Optimal immobilization condition for Amberlite IRA 93 was pH 8.0 and $55^{\circ}C$ yielding 80.6% of immobilization. Activity decay test showed half life of the immobilized enzyme with Amberlite IRA 93 was more than 24 days at $65^{\circ}C$. The carrier was evaluated to be resuable and its result showed the relative immobilization yields were 98.2, 93.3, 90.7 and 87.5%, respectively at second, third, forth and fifth rebinding test of the enzyme on Amberlite IRA 93. Optimal temperature of the immobilized enzyme was slightly lowered and the range widened to $60\sim70^{\circ}C$, while optimal pH moved toward $8.0\sim8.3$ in its isomerization reaction. The trial production result of high fructose corn syrup in pilot scale immobilization showed that one liter of immobilized xylose isomerase (350 IXIU/ml-R) is capable producing about 293l high fructose corn syrup(75% dry substance) in 30 days.