• Asian-Australasian Journal of Animal Sciences
• /
• 제14권3호
• /
• pp.351-357
• /
• 2001

The rapid formation of a cream line cannot be observed in raw goat's milk standing at a low temperature. Although the poor creaming ability of goat's milk has been considered to be due to the small size of milk fat globules and the lack of euglobulin capable of being adsorbed on milk fat globules, there is much left to study. The present work attempted to elucidate a factor for poor creaming ability of goat's milk. The creaming ability of the experimental milks reconstituted from creams and skim milks separated from cow's milk or goat's milk was measured by the volume of the cream layer and the fat content of bottom layer. The polypeptides composition of the P1 the fraction (i.e., the high molecular weight fraction eluted near the void volume obtained by the gel filtration of whey) and milk fat globule membrane prepared from both milks were compared. It was found that the promotion of creaming originated from goat's skim milk was lower than that from cow's skim milk. The P1 fraction in goat's skim milk was less than that in cow's skim milk. The polypeptide (M.W. $4.3{\times}10^4$), found in the P1 fraction of cow's milk was not found in the P1 fraction of goat's milk. It is suggested that the poor creaming ability of goat milk is caused mainly by the difference from cow milk in the amount and the composition of the P1 fraction.

• 한국작물학회지
• /
• 제40권1호
• /
• pp.52-58
• /
• 1995

옥수수의 5-methyltryptophan 저 항성 돌연변이 주（MR1）로부터 anthranilate synthetase（AS）와 tryptophan synthetase（75）의 특성을 분석하였다. 대조구 식물로써 사용한 옥수수 품종 당진의 순계와 저항성주에 있어서 AS와 75의 효소 활성은 5-MT를 포함하지 않은 MS기본배지에서 생장시켰을 때는 차이를 보이지 않았다. 그러나, 25mg/L의 5-MT를 포함한 MS배지에서 생장한 MR1에 있어서 AS의 활성은 대조구보다 2배 높았다. 또한, 4mg/L의 L-tryptophan을 처리 했을때의 AS의 활성은 50％ 저해 되었다. MR1의 조추출물로부터 대조구와 동일한 활성저해율을 나타내기 위해서는 약 4배의 아미노산이 필요하였다. MR1의 75활성은 대조 식물보다 4배가 더 높았다. 그리 하여 tryptophan synthetase B subunit （TSB）를 encoding하는 유전자를 cloning하여 염기배열을 결정 하였다. TSB유전자는 상이한 기관으로부터 cloning된 TSB와 높은 상동성을 보였으며, 모든 발육단계에서 발현하였다. 띠orthern hybridization분석에서 MR1의 TSB발현량은 대조식물보다 높게 나타났다

• 한국잠사곤충학회지
• /
• 제34권1호
• /
• pp.41-48
• /
• 1992

해충의 미생물적 방제를 위하여 Bacillus thuringiensis 살충제(BT제) 개발에 관한 기초자료를 얻고자 3균주의 공시균주인 B. thuringiensis var. kurstaki, B. thuringiensis var. dendrolimus, B. thuringiensis var. aizawai와 3종의 공시충으로 누에, 흰불나방, 배추흰나비의 유충을 이용하고 B. thuringiensis 내독소단백질을 alkalidyddor 또는 공시된 3종의 곤충소화액으로 처리하여 전기영동을 비교하여 아래와 같은 결과를 얻었다. 1. 3균주의 B. thuringiensis에서 생산된 내독소단백질은 alkali 또는 곤충의 소화액 처리에 따라서 변화되는 뚜렷한 차이가 없는 것으로 나타났다. 2. 각 균주의 내독소단백질의 분자량은 B. thuringiensis var. aizawai가 130kDa, B. thuringiensis var. kurstaki가 140kDa, 130kDa, 그리고 B. thuringiensis var. dendrolimus가 140kDa의 부분에 2band로 나타났으며, 이들 내독소단백질은 alkali 용액이나 숙주곤충의 소화액으로 처리시 활성화되어 40-65kDa의 저분자 단백질로 전환되었다. 또한 숙주곤충소화액에 따라 내독소단백질의 활성화 속도에 약간의 차이가 있는 것으로 확인되었다.

• 식물조직배양학회지
• /
• 제22권1호
• /
• pp.15-28
• /
• 1995

보리의 내동성기작에 관여할 것으로 예상되는 반결빙단백질을 분리하기 위해서 저온순화기간 중에 세포밖 공간에 축적되는 단백질을 분리 및 비교하였다. 42일간의 저온처리를 통해 70, 21, 16, 14 KDa의 단백질과 10 KDa 이하의 연속된 크기의 단백질들의 축적이 증가됨을 확인하였다. 이들 단백질들은 3일간의 저온처리에서도 어느 정도 축적되었으나, 42일간의 처리시 그 양에 있어서 더욱 증가됨을 볼 수 있었다. 위 방법으로 얻어진 단백질을 전체 잎조직의 단백질과 비교하여 세포밖 공간의 단백질 추출방법의 정확도를 검증하였다. 또한 호밀의 저온처리시 세포밖 공간에 축적하는 단백질과의 비교를 통해 구성 단백질의 크기에 있어서 차이를 확인하였으나, 호밀과 보리에서 공통적으로 10 KDa 이하의 범위에서 연속적인 크기의 단백질이 축적됨을 볼 수 있었다. 알려진 광어의 반결빙단백질은 크기가 3300에서 33,000 dalton에 이르는 점으로 미루어 보리와 호밀에서 공통적으로 발견되는 10 KDa 이하의 단백질이 반결빙단백질로 작용할 가능성은 매우 높다. Griffith등(1992)은 호밀의 세포밖 공간 단백질중 일부에서 반결빙활성도를 확인하였고 보리와 호밀간의 해당단백질의 전체적인 profile에서의 유사성을 미루어 보리로부터 얻어진 세포밖 공간의 단백질에서 반결빙단백질을 발견할 가능성은 매우 높다.

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

• Journal of Microbiology and Biotechnology
• /
• 제14권6호
• /
• pp.1182-1189
• /
• 2004

Cytosine deaminase (cytosine aminohydrolase, EC 3.5.4.1) stoichiometrically catalyzes the hydrolytic deamination of cytosine and 5-fluorocytosine to uracil and 5-fluorouracil, respectively. The intracellular cytosine deaminase from Chromobacterium violaceum YK 391 was purified to apparent homogeneity with 272.9-fold purification with an overall yield of $13.8\%$. The enzyme consisted of dimeric polypeptides of 63 kDa, and the total molecular mass was calculated to be approximately 126 kDa. Besides cytosine, the enzyme deaminated 5-fluorocytosine, cytidine, 6-azacytosine, and 5-methylcytosine, but not 5-azacytosine. Optimum pH and temperature for the enzyme reaction were 7.5 and $30^{\circ}C$, respectively. The enzyme was stable at pH 6.0 to 8.0, and at 30T for a week. About $70\%$ of the enzyme activity was retained at $60^{\circ}C$ for 5 min. The apparent $K_{m}$ values for cytosine, 5-fluorocytosine, and 5-methylcytosine were calculated to be 0.38 mM, 0.87 mM, and 2.32 mM, respectively. The enzyme activity was strongly inhibited by 1 mM $Hg^{2+},\;Zn^{2+},\;Cu^{2+},\;Pb^{2+},\;and\;Fe^{3+}$, and by o-phenanthroline, $\alpha,\;{\alpha}'$-dipyridyl, p-choromercuribenzoate, N-bromosuccinimide, and cWoramine­T. In addition, the enzyme activity was strongly inhibited by I mM 2-thiouracil, and weakly inhibited by 2-thiocytosine, or 5-azacytosine. Finally, intracellular and extracellular cytosine deaminases from Chromobacterium violaceum YK 391 were found to have a different optimum temperature, apparent $K_{m}$ value, and molecular mass.

• Applied Biological Chemistry
• /
• 제35권6호
• /
• pp.479-484
• /
• 1992

Cheddar 치즈의 숙성기간을 단축시키고 flavor를 증진시킬 목적으로 단백분해력이 있는 Micrococcus sp. LL3를 Cheddar 치즈에 첨가하기 위하여 본 균의 균체생산을 위한 최적배양 온도와 pH 및 배양시간에 따른 casein의 분해형태를 관찰하였다. 본 균주의 생육을 위한 최적 배양온도는 $30^{\circ}C$, 단백분해 효과의 최적 배양온도는 $37^{\circ}C$이었으며, 초기 배양 pH 7.0에서 균의 생육과 단백분해 효과가 가장 좋았다. 배양기간 중 정지기 초기의 세포에서 aminopeptidase의 효소활성이 가장 높았으며, 본 효소의 열 안정성이 높아 $50^{\circ}C$에서 20분간 열처리 후에도 높은 효소활성을 보였다. Casein의 분해속도는 배양 24에서 36시간에서 가장 높았으며, 전기영동을 통한 casein hydrolysis patterns는 ${\alpha}-casein $뿐만 아니라 ${\beta}-casein$도 본 균주에 의해 완전히 분해되었다. 특히 본 균주는 ${\beta}-casein$의 분해력이 더욱 우수하였다.

• Fisheries and Aquatic Sciences
• /
• 제15권4호
• /
• pp.283-291
• /
• 2012

This study examines the optimal fractionation method and conditions for the isolation of endoprotease- and exopeptidase-active fractions from crude extracts of cuttlefish hepatopancreas (HP) using four fractionation methods: ammonium sulfate fractionation (ASF), polyethylene glycol fractionation (PGF), ion exchange chromatography (IEC), and gel filtration chromatography (GFC). Total endoprotease activity highest in the fraction II (concentrate of fractions 34-42; 842.60 U) of GFC, followed by fraction III (40-60% ammonium sulfate fraction; 670.25 U) of ASF, fraction I (concentrate of fractions 8-12; 436.89 U) of IEC, and fraction II (10-20% polyethylene glycol; 307.31 U) of PGF. Total exopeptidase activity of these fractions was highest in fraction II (2,704.70 U) of GFC, fraction III (2,110.50 U) of ASF, fraction III (1,605.60 U) of PGF, and fraction II (concentrate of fractions 38-44; 1,196.22 U) of IEC. These results showed that fraction II of GFC had the highest activity toward both exopeptidase and endoprotease, with exopeptidase activity being 3.21 times higher than of endoprotease. These results suggest cuttlefish HP could be used as a potential source for the extraction of exopeptidase, an enzyme capable of catalyzing the cleavage of N- and C-terminal amino acids in polypeptides, Like endoprotease, the most efficient method for separating exopeptide-active fractions was GFC.

• 약학회지
• /
• 제41권3호
• /
• pp.381-388
• /
• 1997

Metabolism of DWP401, recombinant juman epidermal growth factor, was examined in vivo and in vitro in rats. When $^{125}I$-labeled DWP401 was administered at a dose of 50 ${\mu}g$/kg by i.v. injection. $^{125}I$-labeled DWP401 was rapidly degraded within 30 minytes above 93%. Thin layer chromatography analysis of urine collected for 24 hr after i.v. administration of $^{125}I$-labeled DWP401 showed ohly one spot on a X-ray film which was considered as diiodo-tyrosine. This result suggests tha $^{125}I$-labeled DWP401 was completely digested into free amino acids without any specific intermediate polypeptides. About 42.1% of the administered iodine was recovered in 24 hr. For in vitro degradation study, $^{125}I$-labeled DWP401 was added to plama and tissue homogenates of rats and incubated at $37^{\circ}C$. Almost 98% of the added radioactivity recovered from the protein fraction of the liver, kidey, small intestine, stomach and spleen decreased rapidly. For examplem the recovery rates of $^{125}I$-labeled DWP401 were 58.6, 63.2, 39.9, 52.9 and 66.8% after 4hrs of incubation in respective organ homogenates.

• 대한수의학회지
• /
• 제26권1호
• /
• pp.97-102
• /
• 1986

The prevalence of diarrhea caused by enterotoxigenic Escherichia coli was surveyed on 445 calves in 6 farms which were located in the central part of Korea. The incidence of enterotoxigenic Escherichia coli in calves with diarrhea was investigated by detecting the K99 and F41 antigens from the isolated strains of Escherichia coli The incidence of colibacillosis in calves was 23.3%. Of 238 strains of Escherichia coli isolated from calves with diarrhea, 73 strains(30.6%) were proved possessing the K99 antigen by mannose-resistant hemagglutination(MRHA) using horse red blood cells and 79(33.1%) possessing F41 antigens by MRHA using guinea-pig red blood cells. The minca medium, nutrient broth, tryptic soy broth and brain heart infusion were tested for yield of K99 and F41 pili. The production of pili was greatest in minea medium. The best detachment method of the K99 and F41 pili from the cells was heat treatment for 20 minutes at $60^{\circ}C$ and concentration by precipitation with ammonium sulfate. The purified antigens of K99 and F41 were polypeptides with molecular weights of 18,500 and 29,500, respectively by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.