• 한국식품영양과학회지
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• 제20권5호
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• pp.447-454
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• 1991

쇠고기 단백질 소화에 미치는 마늘의 영향을 검토하기 위해 마늘첨가량, 숙성기간에 따른 효소소화율 및 소화저해물질(Trypsin Inhibitor, TI)의 변화를 실험하였다. 또한 효소 가수분해물의 gel여과 및 가용성부분의 질소량을 정량하여 단백질의 구조 변화를 확인하였으며, 소화율과 아미노산조성 결과를 토대로 예측소화율(Predicted Digestibility, P-dig.), 계산단백효율비(Computed Protein Efficiency Ratio, C-PER) 및 분별계산단백효율비(Discriminant Computed Protein Efficiency Ratio, DC-PER)를 계산하여 단백질 품질을 평가하였다. 쇠고기단백질 소화율은 첨가되는 생마늘량에 약간 영향을 받으나 최적 참가량은 첨가 후의 가열조건에 의하여 결정된다($96{\pm}1^{\circ}C$, 20분 가열시 쇠고기 : 마늘=100g : 12g, 60분 가열시 쇠고기 : 마늘=100g : 3g) 열변성된 쇠고기에 대한 생마늘의 효과는 없고, 날쇠고기육의 소화에는 생마늘의 효과만 인정되며 이미 가열된 쇠고기육의 소화에는 영향을 미치지 못하였다. 날쇠고기를 $4~6^{\circ}C$에서 숙성시켰을 때, 최대의 소화율을 나타내는 시간은 마늘 첨가량에 따라 달라 날쇠고기 100g에 마늘 3g을 첨가할 경우에는(A) 8시간, 12g일 경우에는(B) 12시간이었다. 마늘과 함께 숙성시킨 쇠고기육은 (A)의 경우 $96{\pm}1^{\circ}C$에서 80분, (B)의 경우 20분 정도 가열했을 때 최대소화율을 나타내었다. Four-enzyme으로 가수분해한 숙성시료에는 2,200 dalton 정도의 저분자량의 peptid가 생성되어 소화가 용이함을 확인했으며, 가용성부분도 소화율에 비례하여 증가하였다. 효소소화율에 비례하여 C-PER은 증가하여 C-PER 2.14(날쇠고기)에서 2.50(마늘과 함께 숙성시킨 시료)로 품질이 개선되었다.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2003년도 생물공학의 동향(XII)
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• pp.302-305
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• 2003

It is difficult to obtain sufficient healthy skin for coverage of a wide area of skin wound. In the skin, an additional population of living epithelial cells is located in the outer root sheath (ORS) of hair $follicles.^{1),2)}$ ORS cells should be a good source of epithelium because they are easily obtainable and patients do not have to suffer from scar formation at donor sites. We modified ordinary primary culture technique for the purpose of solving such problem that epithelial cells have a low propagation and easy aging during culture periods. First of all, we improved primary cultivation methods. In the ordinary primary culture, average yield of human ORS cells was $2\;{\times}\;10^3$ cells/follicle by direct incubation with trypsin (0.1%)/EDTA (0.02%) solution for 15 min at $37^{\circ}C$ but we could obtain about $6.5\;{\times}\;10^3$ cells/follicle by two step enzyme digestion method with dispase (1.2 U/ml) and trypsin (0.1%)/EDTA (0.02%) solution. So we could achieve three times higher primary cultured ORS cell yield. Secondly, we could obtain total $2\;{\times}\;10^7$ cells in serum free medium and even more total $6\;{\times}\;10^7$ cells in modified E-medium with mitomycin C-treated feeder cells during 17 days. Using the cultured ORS cells, and we could make bioartificial skin equivalent in vitro and concluded that ORS cells were progenitor cells for skin epithelial cell.

• BMB Reports
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• 제34권2호
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• pp.150-155
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• 2001

The mosquito-larvicidal Cry4B protein from Bacillus thuringiensis subsp. israelensds was expressed in Escherichia coli. Upon activation by trypsin, the 130-kDa protoxin was processed into the 65-kDa active toxin containing two polypeptide fragments of ca. 47 and ca. 20 kDa. These two polypeptides are products of internal cleavages on the exposed loop connecting helices 5 and 6 in the seven-helical bundle domain. PCR-based mutagenesis was employed to introduce an additional cleavage site into the loop connecting helices 3 and 4. A series of amino acid changes were introduced into the targeted loop, resulting in seven mutant protoxins. Upon digestion with trypsin, a group of mutants with arginine introduced into the loop (EPRNQ, EPNRNQ, EPRNP, ESRNP and SSRNP) produced polypeptide products similar to those of the wild type (EPNNQ). When the loop, SSRNP, was expanded by an insertion of either asparagine (NSSRNP) or valine (VSSRNP), an additional cleavage was detected with proteolytic products of 47,12 and 6 kDa. This cleavage was confirmed to be at the introduced arginine residue by N-terminal sequencing. The mosquito larvicidal assay against Aedes aegypti demonstrated a relatively unchanged toxicity for the mutants without cleavage and reduced toxicity for those with an additional cleavage.

• 한국가금학회지
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• 제14권2호
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• pp.89-96
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• 1987

본 연구에서는 가금에 있어 세포유전학적 방법을 이용하여 닭 염색체의 분리방법과 G, C-banding에 의한 분염분석 방법을 재고하여 보다 명확한 염색체의 형태적 양상을 제시하였다. 염색체의 분리는 성장중인 초기배아를 이용하므로써 유사분열 중기상을 쉽게 포악할 수 있었으며, 성장 4-5일째의 배아조직으로부터 염색체의 분리는 다소 불편함이 많았다. 가금에 있어 염색체 분리기술중 가장 중요한 것으로 적합한 sample의 채취와 적절한 중기상의 유도, hypotonic 처리에 따른 뚜렷한 형태의 유도, 도말방법 및 염색의 처리과정이다. 또한 보다 명확한 염색체의 분석을 위하여서는 중기 분열상중 초기 중기 상태의 상을 포착함이 바람직하다. G. C-banding 처리를 위해서는 공히 충분하고도 완전히 건조된 air-dried prepration된 sample을 이용하여야 바람직한 결과를 얻을 수 있으며, slide의 보존시간에 따라 band 양상에 많은 차이를 나타낸다. G-banding 양상에 크게 영향하는 요인으로서는 trypsin의 농도, 처리시간, 처리온도가 주된 작용을 하고, Giemsa solution에 사용하는 buffer의 pH도 크게 작용하는 것으로 나타났다. C-banding에 있어서는 Ba(OJ)$_2$의 농도 처리시간, 처리온도가 큰 영향을 미친 바 다소 짧은 시간의 처리가 바람직한 양상을 보이고, 처리전 HCl 처리로서 세포들의 균일성을 가하고, 처리후 철저한 수세가 좋은 결과를 나타내었다. Band 양상의 보다 정확한 해석을 위해서는 densitometer를 이용하므로써 구체적 양상을 분석할 수 있었다.

• 대한한의학회지
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• 제27권2호
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• pp.103-110
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• 2006

Objectives : This study was conducted to investigate the effects of ginseng saponins and commercial surfactants such as Triton X-100, sodium deoxycholate, and sodium dodecyl sulfate on the gastric enzyme-catalyzed hydrolysis. Methods : Saponins (a surface-active plant component) from fresh ginseng root were extracted to examine its effect on the gastric enzyme-catalyzed hydrolysis. Commercial surfactants such as Triton X-100, sodium deoxycholate, and sodium dodecyl sulfate were also employed in the hydrolysis system to compare their effects with that of the ginseng saponins. The effects of surfactants on the gastric enzyme-catalyzed hydrolysis were measured by using a spectrophotometer. A spectropolarimeter was used to examine the conformational change of enzymes and substrates by the addition of ginseng saponins into the system. Results : Both the tryptic and the peptic digestion of milk casein or eggalbumin were slightly improved with an increase in the amount of ginseng saponins in the system. Triton X-100 showed an effect similar to that of ginseng saponins, while sodium dodecyl sulfate and sodium deoxycholate diminished the hydrolysis. Circular dichroism spectra of enzymes and substrates was significantly changed by the addition of ginseng saponins into the system. Conclusions : These results show that ginseng saponins affect positively the gastric enzyme-catalyzed hydrolysis, and suggest that the digestion of substrates by gastric enzymes is affected by the change of enzyme conformation by ginseng saponins.

• 미생물학회지
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• 제38권2호
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• pp.74-80
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• 2002

단일 탄소원 및 에너지원으로 benzoate를 이용하는 Acinetobacter sp. KS-1에서 분리 정제한 catechol 1,2-dioxygenase (Cl,2O)의 특성과 아미노산 서 열을 분석하였다. Cl,2O는 catechol과 4-methylcatechol에 대해서 효소활성을 나타내었으며, 활성 최적온도는 $35^{\circ}C$이고, 활성 최적 pH는 7.5-9.0의 범위 내에 있었다. 효소활성 저해제로서 은, 수은, 그리고 구리는 Acinetobacter sp. KS-1의 Cl,2O 활성을 억제하였다. SDS-PAGE에 의해 측정된 Cl,2O의 분자량은 약 36 kDa 였으며, N-말단 아미노산 서 열을 분석한 결과, $^{1}MNYQQIDALVKQMNVDTAKG^{20}$로 Acinetobacter radioresistens의 Cl,2O와 95%의 유사성을 보여주었다. In-gel 아미노산 서열 분석을 위하여 trypsin 처리와 peptide mapping을 실시하였다. MALDI-TOF를 이용하여 trypsin으로 처리된 세 개의 peptide flagmen써 분자량을 분석한 결과 966.3 Da, 2081.7 Da, 그리고 1933.8 Da으로 각각 나타났는데, 이는 A. radioresistens의 Cl,2O와 내부 서 열$^{1}SQSDFNLRR^{9}\, ^{1}HGNRPSHVHYFNSAPGYR^{18}\, ^{1}TIEGPLYVAGAPESVGFAR^{19}$ 이 일치하는 것으로 분석되었다. N-말단 서열과 내부 서열을 바탕으로 primer를 제작하여 polymerase chain reaction을 실시하였다.

• 한국축산식품학회지
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• 제34권5호
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• pp.656-664
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• 2014

The purpose of this study was to analyze myosin heavy chain (MHC) isoforms in bovine longissimus thoracis (LT) muscle by liquid chromatography (LC) and mass spectrometry (MS). LT muscles taken from Hanwoo (Korean native cattle) steer (n=3) used to separate myosin bands by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The peptide queries were obtained from the myosin bands by LC-MS/MS analysis following in-gel digestion with trypsin. A total of 33 and 43 queries were identified as common and unique peptides, respectively, of MHC isoforms (individual ions scores >43 indicate identity or extensive homology, p<0.05). MHC-1 (IIx), -2 (IIa), -4 (IIb), and -7 (slow/I) were identified based on the Mowse score (5118, 3951, 2526, and 2541 for MHC-1, -2, -4, and -7, respectively). However, more analysis is needed to confirm the expression of MHC-4 in bovine LT muscle because any query identified as a unique peptide of MHC-4 was not found. The queries that were identified as unique peptides could be used as peptide markers to confirm MHC-1 (14 queries), -2 (8 queries), and -7 (21 queries) in bovine LT muscle; no query identified as a unique peptide of MHC-4 was found. LC-MS/MS analysis is a useful approach to study MHC isoforms at the protein level.

• Asian-Australasian Journal of Animal Sciences
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• 제9권6호
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• pp.701-703
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• 1996

The influence of phenylalanine (Phe) in the medium on protein synthesis of chicken embryo fibroblasts (CEF) was examined. CEF was derived from 9-d-old embryos by trypsin-EDTA digestion. To examine the deficiency of Phe in the medium, CEF was cultured in Dulbecco's modified Eagle's medium (DMEM) with or without Phe. CEF was also cultured in Dulbecco's phosphate buffered saline (PBS ($Ca^{2+}$, $Mg^{2+}$)) with or without $400{\mu}m$ Phe in order to examine the effect of Phe supplementation. All media were supplemented with 10% (v/v) fetal calf serum. After incubation for 6, 30 and 54 h, protein synthesis was measured by the incorporation of L-[2, $6-^{3}H$] Phe into CEF for further 18 h. Protein synthesis of CEF cultured in DMEM was higher than that in PBS ($Ca^{2+}$, $Mg^{2+}$). High specific radioactivity of Phe due to the low concentration of Phe in the medium resulted in the apparent increase in protein synthesis of CEF. Protein synthesis cultured in PBS ($Ca^{2+}$, $Mg^{2+}$) with Phe did not increase during 72 h of cell culture.

• 한국동물위생학회지
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• 제15권2호
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• pp.109-120
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• 1992

330 Samples of the slaughtered cattle heart muscle were collected from the abattoirs of five regions in Kangwon - do to reveal the frequency of sarcocystis infections during January through December in 1991. The samples were inspected for bradyzoites by the trypsin digestion technique and the possitive samples were fed to dogs and cats for the detection of sporocysts shed in the feces. The results obtained were summarized as follows : 1. The infection rate of bovine Sarcocystis investigated from 330 samples was 43.6%. 2. It revealed that the infection rate of Sarrocystis increased gradully with the advance in the age, 14.5% in below two years, 26.1% in the three years, 30% in four years, 54.7% in five years, 74.4% in six years, 90% in seven years and 100% in older than eight years. 3. The cyst walls detected out from the heart muscles were less than l${\mu}{\textrm}{m}$ in thickness and the size of bradyzoites were $11.8{\times}2.8{\mu}m$ in average. 4. The size of sporocysts shed in the feces of dogs were $15.8{\times}9.8{\mu}m$ in average and the prepatent periods ranged from 12 to 16days. 5. Sarcorystis found in the bovine heart muscles were identified as Sascocystis cruzi ( Hasselman, 1923) , wenyon, 1926.

• 한국수산과학회지
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• 제44권5호
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• pp.456-463
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• 2011

We attempted to isolate a collagenase-1 inhibitory peptide from skate Dipturus chilensis skin protein. The protein from skate skin was digested by various enzymes (alcalase, ${\alpha}$-chymotrypsin, neutrase, papain, pepsin, and trypsin) to produce a collagenase-1 inhibitory peptide. The collagenase-1 inhibitory activity of the peptides obtained was measured by gelatin digestion assay. Among the six hydrolysates, pepsin hydrolysate exhibited the highest collagenase-1 inhibitory activity. The peptide showing strong collagenase-1 inhibitory activity was purified by Sephadex G-25 gel chromatography and HPLC using an octadecylsilyls (ODS) column. The amino acid sequence of purified collagenase-1 inhibitory peptide was identified to be Asn-Leu-Asp-Val -Leu-Glu-Val-Phe (961 Da) by quadrupole time of flight (Q-TOF) and electrospray ionization mass spectrometry (ESI-MS) mass spectroscopy. The $IC_{50}$ value of purified peptide was 87.0 ${\mu}M$. Moreover, the peptide did not exhibit cytotoxic effects on human dermal fibroblast cell lines.