• Asian-Australasian Journal of Animal Sciences
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• 제24권12호
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• pp.1718-1728
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• 2011

Two experiments were conducted to investigate the in vitro ability of keratinase to hydrolyze soybean glycinin and ${\beta}$-conglycinin and to evaluate the in vivo effects of keratinase when included in corn-soybean diets with different levels of crude protein and fed to nursery pigs. In experiment 1, a saturated keratinase solution (1 ml) was added to two blank controls of either glycinin or ${\beta}$-conglycinin resulting in the hydrolysis of 94.74% glycinin and 88.89% ${\beta}$-conglycinin. In experiment 2, 190 pigs (8.3${\pm}$0.63 kg BW) were allotted to one of four treatments in a 2${\times}$2 factorial arrangement on the basis of body weight, and sex was balanced among the pens. The effects of crude protein (19 vs. 22%) and keratinase (0 vs. 0.05%) were studied. Each treatment was applied to six pens with seven (two pens) or eight pigs per pen. Pigs were fed the experimental diets for 21 d. Weight gain and feed conversion ratio were improved (p<0.05) with keratinase supplementation while feed intake was reduced (p<0.05). Keratinase supplementation increased (p<0.05) the apparent total tract digestibility of dry matter, energy, crude protein and phosphorus. Keratinase supplementation also increased n-butyric acid in the cecum and colon, lactobacilli and total anaerobe counts in the colon as well as the ratio of villus height to crypt depth in the ileum. Additionally, fecal score, ammonia nitrogen and branch chain volatile fatty acids in the colon, E. coli and total aerobe counts in the colon, crypt depth in the jejunum and ileum as well as serum interleukin-1 and interleukin-6 concentrations were also decreased (p<0.05) by keratinase supplementation. A reduction in dietary crude protein decreased (p<0.05) colon ammonia nitrogen concentration and cecal propionic acid and branch chain volatile fatty acid concentrations. In addition, cecal E. coli counts, colon total anaerobe counts, ileal crypt depth, and serum interleukin-1 and interleukin-6 concentrations were also decreased (p<0.05) with the reduction of dietary crude protein. With the exception of fecal scores, there were no significant interactions between crude protein and keratinase. This study provides evidence that dietary keratinase supplementation improved nursery pig performance by improving intestinal morphology and ecology, thus improving nutrient digestibility and alleviating the inflammatory response.

• 한국식품영양과학회지
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• 제35권4호
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• pp.470-475
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• 2006

국내산 콩 품종 3종과 미국산 콩 2종(non-GMO 및 GMO glyphosate-tolerant HS2906)에 대하여 단백질함량과 분포의 차이를 아미노산 분석, 총 질소량, PAGE/densitometry법에 의 해 분석하였다. 아미노산 구성과 총 질소량에서 품종간 유의적인 차이는 없었다. 한편 단백질 추출에서 SDS/buffer 추출법이 더 효과적이었다. 총 단백질함량(380.2-423.9 mg/g)과 단백질 분포는 품종간 유사하였으나. 상세 PACE(12.5% normal gel) 결과 재래종 콩(WS82)에 비해 제초제 저항성 GMO콩(HS2906)에서 beta conglycinin(55kDa) 양이 낮게 나타났으며, 25 kDa 단백질의 경우 미국산 콩에서는 관찰되지 못한 반면에 국내산 콩 품종에서는 뚜렷하게 관찰되었으며, ${\beta}$-conglycinin은 미국산 제초제 저항성 GMO콩과 국내산 황금콩에서 상대적으로 낮은 것으로 관찰되었다. 이와 같은 결과는 normal PAGE/densitometry 분석법은 콩시료 분석에 유용하게 활용될 수 있음을 보여주었다.

• 한국식품영양과학회:학술대회논문집
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• 한국식품영양과학회 2001년도 International Symposium on Food,Nutrition and Health for 21st Century
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• pp.69-73
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• 2001

Two opioid peptides, YPLDL and YPLDLF, were isolated from enzymatic digests of spinach ribulose-1, 5-bisphosphate carboxylase/oxygenase (RuBisCO) and named rubiscolin-5 and -6, respectively. These peptides were selective for delta-receptor and the latter was about 3 times more potent than the former. After oral administration in mice at the dose of 100 mg/kg, rubiscolin-6 showed analgesic activity in tail pinch test. It also stimutated learning performance at the same dose in passive avoidance experiment using step-through apparatus. An immunostimulating peptide, MITLAIPVNKPGR, was isolated from a trypsin digest of soybean protein and named soymetide. Immunostimulating activy of soymetide was mediated by fMLP receptor. Interestingly, after oral administration in rats at a dose of 300 mg/kg (po.), soymetide-4 (MITL) protected alopecia (hair-loss) induced by etoposide, a cancer chemotherapy agent. Stimulation of IL-1 release by the peptide was involved in the mechanism. Ovokinin(2-7), RADHPF, is a vasorelaxing peptide released from ovalbumin by the action of chymotrypsin. It lowered blood pressure of spontaneously hypersensive rats (SHR) after oral administration at a dose of 10 mg/kg. RPLKPW, which was designed by replacing 4 amino acid residues in ovokinin(2-7), exhibited hypotensive activity at a dose of 0.1 mg/kg (po.). This peptides was introduced into 3 homologous sites in soybean beta-conglycinin alpha' subunit by site-directed mutagenesis of the cDNA and expressed in E. coli. The minimum effective dose for hypotensive activity of the genetically modified beta-conglycinin alpha' subunit was 10 mg/kg (po.), which is about 1/200 that of ovalbumin.

• 한국육종학회지
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• 제42권1호
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• pp.35-39
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• 2010

Soybean proteins are widely used for human and animal feed in the world. ${\beta}$-conglycinin protein exhibits poor nutritional and food processing properties and Kunitz trypsin inhibitor (KTI) protein is a main anti-nutritional factor in soybean seed. The objective of this research was to identify the inheritance of $cgy_1$ gene and ti gene for the improvement of soybean cultivar with no KTI proteins and low amount of ${\beta}$-conglycinin. $F_2$ population was made by crossing between "Gaechuck2ho" (${\alpha}^{\prime}$-subunit present $Cgy_1Cgy_1$, KTI protein absent titi) and PI506876 (${\alpha}^{\prime}$-subunit absent $cgy_1cgy_1$, KTI protein present TiTi) parent. A total of 434 $F_2$ seeds were obtained and analyzed for the segregation of ${\alpha}^{\prime}$-subunit protein and KTI protein using SDS-PAGE. The segregation ratio of 3 : 1 for $Cgy_1$ locus (310 $Cgy_1$_ : 124 $cgy_1cgy_1$) and Ti locus (339 Ti_ : 95 titi) were observed. Segregation ratios of 9 : 3 : 3 : 1 (241 $Cgy_1$_Ti_: 69 $Cgy_1$_titi: 98 $cgy_1cgy_1$Ti_: 26 $cgy_1cgy_1titi$) between $Cgy_1$ gene and Ti gene in $F_2$ seeds were also observed (${\chi}^2= 5.367$, P = 0.10 - 0.20). This data showed that $Cgy_1$ gene was inherited independently with the Ti gene in soybean. These results will be useful in breeding program for selecting the line that does not exhibit or lacks both ${\alpha}^{\prime}$-subunit protein and KTI protein in soybean.

• 한국식물학회:학술대회논문집
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• 한국식물학회 1987년도 식물생명공학 심포지움 논문집 Proceedings of Symposia on Plant Biotechnology
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• pp.283-307
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• 1987

Glycinin and $\beta$-conglycinin are the most abundant storage protein in soybean. These proteins are known to be synthesized predominantly during germination and cell expansion phase of seed development for short period, and synthesized not in other tissues. Genes encoding these storage proteins are useful system to study the mechanism of development stage and tissue specific gene expression in eukaryotes, especially plants, at the molecular level. The cDNA and genomic clones coding for glycinin have been isolated and regulation mechanism of the gene expression has been studied. Initially, development and tissue-specific expression of the glycinin gene is regulated at the level of transcription. Post-transcriptional processing is also responsible for delayed accumulation of the mRNA. Translational control of the storage protein gene has not been reported. Post-translational modification is another strategic point to regulate the expression of the gene. It is possible to identify positive and/or negative reguratory clements in vivo by producing transgenic plants agter gene manipulation. Elucidation of activation and repression mechanism of soybean storage protein genes will contribute to the understanding of the other plant and eukaryotic genes at molecular level.

• Plant Resources
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• 제1권2호
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• pp.113-120
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• 1998

Soybean is an important crop because its seed has very high protein relative to others. The quality of soy protein is limited by the concentration of the sulfur-containing amino acids in the amino acid profile. Among the supply of various forms of 0.4mM sulfur as S nutrition during seed fill. only 0.4mM L-methionine can inhibit ${\beta}$-subunit synthesis completely and produce the highest glycinin-containing seeds. Compared to 0.4mM sulfate control, seeds supplied by 0.4mM L-methionine have lower ${\alpha}$-, no ${\beta}$-subunit, and highly increased glycinin without altering total protein concentration. Supply of 0.2mM cystine (0.4mM S) did not affect the accumulative pattern of seed storage protein (SSP) subunits. In the supply of L-methionine, 0.2mM treatment showed higher glycinin in seeds but 0.05mM resulted in lower glycinin than tile sulfate control. The relative abundance of ${\alpha}^`$-subunit was not altered by any N or S nutrition. Under 5mM nitrogen, protein concentration was increased about 3-5% by substituting ammonia for nitrate during seed fill independent of nutrition. The increase resulted in the only increase of 7S protein, mainly ${\beta}$-subunit. Our data suggest that the regulatory system of SSP genes responds to the balance between N and S assimilates supplied from mother plant. and controls the di fferential synthesis of their subunits for the maximum protein accumulation in developing soybean seed.

• Journal of Applied Biological Chemistry
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• 제51권3호
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• pp.88-94
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• 2008

Three different protein extraction methods-phenol extraction, trichloroacetic acid (TCA) precipitation, and desalting/TCA precipitation-were compared to determine the optimal reproducible high resolution 2-dimensional (2-D) electrophoresis for each chungkugjang, doenjang, and kochujang samples. The soluble proteins from Chungkugjang extracted by phenol were separated with high reproducibility and resolution, and gained 1.75- to 3-fold more protein spots on 2-D gel than those from the other methods. On the contrary, the extracted proteins from doenjang and kochujang treated by desalting/TCA precipitation method showed about 1.5- to 3.3-fold more protein spots on 2-D gel. Using the established methods, the changes in the protein profiles of the fermented soy foods were monitored during the fermentation period by 2-DE. One of the major proteins in soy, $\beta$-conglycinin $\alpha$-subuint, and some proteins with unknown functions were localized on 2-D gel as the protease-resistant proteins throughout the fermentation period of doenjang. Changes in the protein profile monitored by the established methods can provide basic information on unfolding the mechanisms of the generation of biofunctional activity in the fermented soy foods.

• 한국작물학회지
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• 제50권1호
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• pp.32-38
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• 2005

A long-term study initiated in 1989 at San-born Field, Columbia, Missouri, was designed to evaluate the affect of environmental factors, nitrogen application, and crop rotation on soybean (Glycine max [L.] Merr.) seed composition. Soybeans were grown as part of a four- year rotation which included corn (Zea maize L.), wheat (Triticum aestivum L.), and red clover (Trifolium pratense L.). Results from soil tests made prior to initiation of the study and subsequently every five years, were used to calculate application rates of nitrogen, phosphorus, and potassium necessary for target yield of pursuant crops. In the experimental design, nitrogen was applied to one-half of the plot on which the non-leguminous crop, either corn or wheat was grown. Analysis of soybean seed by near infrared reflectance spectroscopy collected over an 11-year period revealed a linear increase in protein and decrease in oil content. Application of nitrogen fertilizer to non-leguminous crops did not have an apparent effect on total protein or oil content of subsequent soybean crop. Analysis of soybean seed proteins by sodium dodecyl sulfate polyacrylamide gel electrophoresis in conjunction with computer­assisted densitometry revealed subtle changes in the accumulation of seed proteins. Immunoblot analysis using antibodies raised against the $\beta-subunit$ of $\beta-conglycinin$ showed a gradual increase in the accumulation of the 7S components during successive years of the experiment. A linear increase in temperature and decrease in rainfall was observed from the onset of data· collection. Higher temperatures during the growing season have been linked to increased protein and diminished oil content of soybean, thus changes observed in this study are possibly related to climatic conditions. However, crop rotation and subsequent changes in soil ecology may contribute to these observed changes in the seed composition.

• Applied Biological Chemistry
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• 제38권5호
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• pp.387-392
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• 1995

대두 glycinin 유전자의 조직 특이적이고 분화 발달 특이적인 발현 조절 메카니즘을 연구하기 위하여 Gy2 유전자의 5' upstream 부위 염기서열을 조사한 결과, glycinin 유전자의 발현을 조절하는 인자로 여겨지는 여러 가지 조절 인자들을 발견하였다. 진핵세포 유전자에 공통적으로 존재하는 TATA box와 AGGA box가 존재하고, 종자 저장 단백질에서 공통적으로 발견되는 embryo factor binding sequence, RY repeat CACA sequence, ${\beta}$-conglycinin enhancer 와 유사한 sequence 등이 발견되었다. 이러한 조절 요소들이 Gy2 유전자의 발현 조절에 미치는 영향을 알아보기 위해 Gy2유전자의 5' upstream부위를 Exo III nuclease와 여러가지 제한효소를 이용하여 일련의 deletion mutants를 제조한 후 GUS 유전자와 결합시켰다. 이들 여러가지 chimeric constructs를 대두 원형질체에 전입하고 원형질체로부터 추출물을 분리하여 GUS 활성을 조사한 결과, $-28l{\sim}-223$ 혹은 $-l70{\sim}-122$ 부위를 포함하였을 경우 활성이 감소하였고, $-223{\sim}-170$ 혹은 $-l22{\sim}-16$ 부위를 포함하였을 경우 활성이 높게 나타났다. 이러한 Gy2 유전자의 이중적인 발현 양상은 glycinin 유전자의 발현조절에 음성 조절 요소와 양성 조절 요소가 관여하고있다는 사실을 제시해 주고 있다. 또한 이들 여러가지 chimeric constructs로 형질 전환된 담배의 종자와 잎에서 GUS활성을 조사한 결과, CaMV promoter를 포함하는 chimeric construct는 종자와 잎에서 모두 활성을 나타냈으나, Gy2 Promoter를 포함하는 chimeric constructs는 종자에서만 GUS 활성을 나타내고 잎에서는 활성이 나타나지 않는 조직 특이적인 발현 양상을 나타내었다.

• 한국식품조리과학회지
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• 제13권1호
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• pp.78-85
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• 1997

본 연구에서는 응고제의 종류를 달리하여 제조한 두부의 텍스쳐 특성을 알아보기 위해 기계적 측진과 관능검사를 실시하였고, 두부 제조시 부산물로 생기는 두부순물의 일반성분, 올리고당, 아미노산을 분석하여 비교하였으며, 콩분말, 두유, 두부순물의 단백질 pattern을 전기 영동을 실시하여 알아보았다. 1. 두부의 기계적 견고성, 응집성, 탄력성, 검성은 수분함량이 적었던 염화칼슘두부와 염화마그네슘두부가 황산칼슘두부, 글루코노델타락톤두부보다 컸다. 2. 두부의 관능검사 결과 텍스쳐는 황산칼슘두부, 글루코노델타락톤두부가 수분함량이 많아 부드럽고 매끈하다고 평가되었으나 글루코노델타락톤두부는 신맛을 강하게 나타내었다. 3. 동결건조한 글루코노델타락톤두부에서 나온 두부순물에는 상데적으로 arginine이 가장 많았으나, 나머지 3종류의 주부순물의 아미노산 중에는 glutamic acid가 가장 많았다. 4. 두부 순물에 함유된 주요 당은 fructose, glucose, sucrose, raffinose, stachyose였으며 당의 총 함량은 12-13 g/l 내외로 순물간에 차이가 별로 없었다. 5. 전기영동결과 콩분말과 두유에서는 콩단백진의 주요 획분인 glycinin과 $\beta$-conglycinin이 나타났으나 두부 순물에서는 저분자량의 band 만이 나타났다.