• BMB Reports
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• v.28 no.5
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• pp.375-381
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• 1995

Two forms of glucoamylase (GI and GII) from starch-grown Lipomyces kononenkoae CBS 5608 mutant were purified to apparent homogeneity by means of ultrafiltration, Sephacryl S-200 gel filtration and DEAE Sephadex A-50 chromatography. The apparent molecular weight was calculated as ca. 150 kDa for GI and ca. 128 kDa for GII, respectively. Both enzymes were glycoproteins with isoelectric points of 5.6 (GI) and 5.4 (GII). They had a pH optimun of 4.5 and were stable from pH 5 to 8. The temperature optimum for both enzymes was $60^{\circ}C$, but they were rapidly inactivated above $70^{\circ}C$. The $K_m$ values toward starch were estimated to be 6.57 mg per ml for GI and 4.52 mg per ml for GII, and the $V_{max}$ values were 16.28 ${\mu}M$ per mg for GI and 32.25 ${\mu}M$ per mg for GII, respectively. The $K_m$ and $V_{max}$ values of GII for ${\alpha}-$ or ${\beta}-cyclodextrin$ were estimated to be 0.15 mg per ml and 2.0 mg per ml, respectively ($K_m$) and 1.02 ${\mu}M$ per mg or 1.02 ${\mu}M$ per mg, respectively ($V_{max}$). Neither enzyme exhibited pullulanase activity but they released only glucose from starch or cyclodextrin. Amino acid analysis indicated that both glucoamylases were enriched in proline and acid amino acids. Glucoamylase GII strongly cross-reacted with a monoclonal antibody raised against GI enzymes, and the two enzymes shared very similar amino acid composition. Western blot analysis indicated that L. kononenkoae CBS 5608 mutant produced two forms of glucoamylase on starch, and that synthesis of them was subject to glucose repression.

• Microbiology and Biotechnology Letters
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• v.45 no.3
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• pp.226-235
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• 2017

Carotenoids such as phytoene, lycopene, and ${\beta}-carotene$ are used as food colorants, animal feed supplements, and for human nutrition and cosmetic purposes. Previously, we reported the isolation of a novel marine bacterium, Kocuria gwangalliensis, which produces a pink-orange pigment. Phytoene desaturase (CrtI), encoded by the gene crtI, catalyzes lycopene formation from phytoene and is an essential enzyme in the early steps of carotenoid biosynthesis. CrtI is one of the key enzymes regulating carotenoid biosynthesis and has been implicated as a rate-limiting enzyme of the pathway in various carotenoid synthesizing organisms. Here, we report the cloning of the crtI gene responsible for lycopene biosynthesis from K. gwangalliensis. The gene consisted of 1,584 bases encoding 527 amino acid residues. The nucleotide sequence of the crtI gene was compared with that of other species, including Kocuria rhizophila and Myxococcus xanthus, and was found to be well conserved during evolution. An expression plasmid containing the crtI gene was constructed (pCcrt1), and Escherichia coli cells were transformed with this plasmid to produce a recombinant protein of approximately 57 kDa, corresponding to the molecular weight of phytoene desaturase. Lycopene biosynthesis was confirmed when the plasmid pCcrtI was co-transformed into E. coli containing the plasmid pRScrtEB carrying the crtE and crtB genes required for lycopene biosynthesis. The results from this study will provide valuable information on the primary structure of K. gwangalliensis CrtI at the molecular level.

• Fisheries and Aquatic Sciences
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• v.18 no.3
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• pp.311-316
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• 2015

Ten specimens of Brama dussumieri (family Bramidae) were collected from waters off Jeju Island, Busan, and Gangneung, Korea, during 2013-2014. The specimens were characterized by having 58-64 lateral line scales and 13-15 gill rakers. An analysis of 567 base pair sequences of mitochondrial DNA cytochrome c oxidase subunit I showed that sequences in our ten specimens are concordant with those of B. dussumieri from the USA, India, and Japan, although with slight differences (genetic distance = 0.000-0.018). Brama dussumieri was distinguished from the most similar species, Brama japonica, by the number of lateral line scales (57-65 in B. dussumieri vs. 65-75 in B. japonica) and the number of gill rakers (13-15 in B. dussumieri vs. 17-20 in B. japonica). We propose the new Korean name "Wae-sae-da-rae" for B. dussumieri in Korea.

• Journal of Applied Biological Chemistry
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• v.57 no.1
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• pp.61-63
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• 2014

Aged black garlic (ABG) was extracted with 20% ethanol and water (crude extracts) and fractionated into three categories (>10, 3-10, and <3 kDa). The effect of crude extract supplements on anti-My-10 hybridoma cell growth and IgG1 antibody production was investigated in suspension culture with a chemically defined protein-free medium. We observed that supplementation of ABG to the cell culture medium stimulated anti-My-10 hybridoma cell growth and production of IgG1 antibody, particularly with fractionated ABG of low molecular weight. The stimulation depended upon the concentration and the size of the fractionated ABG. We also found that the growth-promoting activity was not correlated with high antibody production. These results suggest that fractionated ABG is a novel and promising alternative as an animal cell culture supplement.

• Journal of the Korean Society of Food Culture
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• v.34 no.6
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• pp.785-792
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• 2019

This study examined the changes in the protein and mineral composition of Gryllus bimaculatus fermented with Bacillus substilis and the mycelia of Basidiomycetes. Normal Gryllus bimaculatus (S) and experimental group data obtained after an inoculation of Bacillus substilis (SC) (KACC 19623), Pleurotus eryngii (SP) and Cordyceps millitaris (SC) were compared. The crude protein content of the Gryllus bimaculatus (control) was 75.48%, but it decreased to 64.55, 54.32, and 63.53% after fermentation with SB, SP and SC, respectively (p<0.05). An analysis of the organic elements showed that the contents of the carbon and nitrogen sources were also reduced after fermentation, and the most significant decrease was observed after fermentation with SP. In SDS-PAGE, a 120 kDa and a 48 kDa protein of Gryllus bimaculatus were found. On the other hand, protein bands faded after fermentation with SP and SC, respectively. Moreover, no visible band was observed after fermentation with SB. According to amino acid analysis, the total free amino acid content increased 3.84 and 1.74 times after fermentation with SB and SP, respectively, compared to the corresponding baseline data. In contrast, it decreased by 0.52 times after fermentation with SC. Among the essential amino acids found in crickets fermented with SB, the valine and isoleucine content was 3.57 and 2.64 times higher, respectively, than the recommended daily amount of essential amino acids.

• Transactions of the Korean Society of Mechanical Engineers B
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• v.34 no.5
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• pp.491-498
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• 2010

Humidification of PEM fuel cells is necessary for enhancing their performance and lifetime. In this study, a humidification system was designed and tested; the system includes an air-supply tube (inner diameter: 75 mm) through which a nozzle can be directly inserted and a cyclonic separator for the removal of water droplets. Three types of nozzles were employed to study the influence of injection pressure, air flow rate, and spray direction on the humidification performance. To evaluate the humidification performance, the concept of humidification efficiency was defined. In the absence of an external heat source, latent heat for evaporation will be supplied by the own enthalpies of water and air. Thus, the amount of water sprayed from the nozzle is the most critical factor affecting the humidification efficiency. Water droplets were efficiently removed by a cyclonic separator, but re-entrainment occurred at high air flow rates. The absolute humidity and humidification efficiency were $21.29\;kJ/kg_{da}$ and 86.57%, respectively, under the following conditions: nozzle type PJ24; spray direction angle $90^{\circ}$; injection pressure 1200 kPa; air flow rate 6000 Nlpm.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.32 no.1
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• pp.75-82
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• 1999

The hydrolysate of desalinated tuna boiled extract (TBE) were prepared by continuous hydrolysis of TBE using a membrane reactor. TBE and tuna boiled extract hydrolysate (TBEH) were isolated depending on molecular weights. The major molecular weight distributions of TBEH-l0K, TBEH-5K and TBEH-lK were 9,800Da, 3,000Da and 990Da, respectively. The amounts of nucleotides and their related compounds of TBE were 3.47 $\mu$mole/g AMP, 23.75 $\mu$mole/g IMP, 9.07 $\mu$mole/g inosine and 1.89 $\mu$mole/g hypoxanthine. Total content of amino acids having desirable taste (glycine, glutamic acid, alanine, proline, aspartic acid, serine) was about $63\%$ of total amino acid from TBE and about $62\%$ from TBEH. The natural seasoninings were prepared with TBE and TBEH. From the results of sensory evaluations, complex seasoning containing TBEH-1K was almost equal to the shellfish complex seasoning obtained from the market. The mixed sauce which was made by mixing of $50\%$ TBEH sauce and $50\%$ fermented soy sauce was similar to the tradition soybean sauce in product quality and it showed the possibility to be used for the substitute product for acid hydrolyzed soysauce.

• Research in Plant Disease
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• v.12 no.3
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• pp.226-230
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• 2006

Cucumber mosaic virus(CMV) was occurred on red pepper showing vein chlorosis or vein necrosis with the incidence rate of 52% from 62 specimens collected in natural fields. Among 32 samples infected with CMV, the specimens of 22 red pepper leaves showing vein chlorosis were infected singly with CMV-VCH. CMV-VCH induced vein chlorosis on the inoculated leaves, and vein banding and vein necrosis on the upper leaves of Nicotiana glutinosa, and then killed after showing bud necrosis. The typical symptoms of vein banding, malformation and blister were produced on the upper leaves of Nicotiana benthamiana and N. tabacum 'Ky-57' without symptoms on the inoculated leaves. The commercial cultivars of 'Bugang', 'Manitta' and 'Gwariput' were shown the typical symptom of vein chlorosis by the mechanical inoculation of CMV-VCH. CMV-VCH was detected specifically by RT-PCR. Virus particles of CMV-VCH were isometric shape having 30 nm diameter. Ultraviolet absorption of purified CMV-VCH was maximum at 260 nm and minimum at 242 nm. The ratio of A260/A280 was 1.71. CMV-VCH had the single nucleo-protein having the molecular weight of 24.5 kDa.

• BMB Reports
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• v.38 no.1
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• pp.49-57
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• 2005

Major Royal Jelly Protein cDNAs of Apis cerana (AcMRJP) were cloned and characterized. The open reading frames (ORFs) of the AcMRJP1 and AcMRJP2 genes were 1302 and 1392 nucleotides, encoding 433 and 463 amino acid residues, respectively. The sequence divergences between AcMRJP1 and AcMRJP2 and their corresponding protein families in A. mellifera were 0.0618 and 0.0934 at the nucleotide level and 0.0912 and 0.1438 at the protein level, respectively. Phylogenetic analysis supports the orthologous similarity between these proteins. The deduced amino acids indicated high essential amino acid contents of AcMRJP1 and AcMRJP2 (47.5 and 44.8%, respectively). The genomic organization of both AcMRJP1 and AcMRJP2 was determined. Both the AcMRJP1 (3663 bp) and AcMRJP2 (3963 bp) genes contained six exons and five introns, where all boundaries conformed to the GT/AG rule. AcMRJP1 and AcMRJP2 cDNAs were cloned into pET17b, and both the recombinant (r) AcMRJP1 (47.9 kDa) and rAcMRJP2 (51.7 kDa) were expressed in the insoluble form. Western blot analysis and N-terminal sequencing of the solubilized proteins revealed successful expression of rAcMRJP1 and rAcMRJP2 in vitro. The yields of the purified rAcMRJP1 and rAcMRJP2 were approximately 20 and 8mg protein per liter of the flask culture, respectively.

• Journal of Pharmaceutical Investigation
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• v.40 no.3
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• pp.175-180
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• 2010

Albumin-modification has been viewed as one of the most effective ways of extending the short in vivo lifetimes of peptide drugs by delaying glomerular filtration. In this study, we describe a new type 2 anti-diabetic exendin-4 (Ex4) peptide derivative with significant binding ability to human serum albumin (HSA). This exendin-4 derivative consists of a 4-branched polyethylene glycol $(PEG)_{5k}$ (Mw: 20 kDa) modified with three stearylamines ($C_{18}-NH_2$) and one exendin-4 on its branches. PEG and stearylamine were selected to provide functionality to increase molecular size and bind to albumin, respectively. This derivative ($3C_{18}-4PEG_{5k}$-Ex4) was shown to have larger molecular size (Ca. 152 kDa) than actual (25.0 kDa) when subjected to size-exclusion chromatography, and the fluorescein-tagged $3C_{18}-4PEG_{5k}$-Ex4 displayed significant binding to the HSA-immobilized Sepharose CL-4B resin using confocal laser scanning microscopy. Furthermore, $3C_{18}-4PEG_{5k}$-Ex4 was found to have acceptable anti-hyperglycemic efficacy via three consecutive oral glucose tolerance testings (OGTT) in fasted type 2 diabetic db/db mice. The $HD_{total}$ value ($57.6{\pm}12.3%$) of $3C_{18}-4PEG_{5k}$-Ex4 at a 50 nmol/kg dose was 2-fold greater than that ($31.0{\pm}8.7%$) of native exendin-4 in non-fasted db/db mice. Especially, the blood glucose levels in the mice group treated with $3C_{18}-4PEG_{5k}$-Ex4 did not rebound to ~150 mg/dL until 24 h after the injection, which obviously shows the extended hypoglycemia. We believe that this derivative has great pharmaceutical potential as a novel long-acting type 2 anti-diabetic injection treatment.