• Journal of Microbiology and Biotechnology
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• 제22권11호
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• pp.1532-1539
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• 2012

The ${\alpha}$-galactosidase-coding gene agaAJB13 was cloned from Sphingomonas sp. JB13 showing 16S rDNA (1,343 bp) identities of ${\leq}97.2%$ with other identified Sphingomonas strains. agaAJB13 (2,217 bp; 64.9% GC content) encodes a 738-residue polypeptide (AgaAJB13) with a calculated mass of 82.3 kDa. AgaAJB13 showed the highest identity of 61.4% with the putative glycosyl hydrolase family 36 ${\alpha}$-galactosidase from Granulicella mallensis MP5ACTX8 (EFI56085). AgaAJB13 also showed <37% identities with reported protease-resistant or Sphingomonas ${\alpha}$-galactosidases. A sequence analysis revealed different catalytic motifs between reported Sphingomonas ${\alpha}$-galactosidases (KXD and RXXXD) and AgaAJB13 (KWD and SDXXDXXXR). Recombinant AgaAJB13 (rAgaAJB13) was expressed in Escherichia coli BL21 (DE3). The purified rAgaAJB13 was characterized using p-nitrophenyl-${\alpha}$-D-galactopyranoside as the substrate and showed an apparent optimum at pH 5.0 and $60^{\circ}C$ and strong resistance to trypsin and proteinase K digestion. Compared with reported proteaseresistant ${\alpha}$-galactosidases showing thermolability at $50^{\circ}C$ or $60^{\circ}C$ and specific activities of <71 U/mg with or without protease treatments, rAgaAJB13 exhibited a better thermal stability (half-life of >60 min at $60^{\circ}C$) and higher specific activities (225.0-256.5 U/mg). These sequence and enzymatic properties suggest AgaAJB13 is the first identified and characterized Sphingomonas ${\alpha}$-galactosidase, and shows novel protease resistance with a potential value for basic research and industrial applications.

• 한국작물학회지
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• 제27권1호
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• pp.60-65
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• 1982

맥류의 한발저항성 반응과 정도 그리고 저항생리적 기작을 구명하기 위하여 발아후 10일된 유묘기(본엽 3매시)에 8일간 단수처리를 하여 질산환원효소와 단백질분해효소의 활성변화 그리고 proline축적을 조사하였던 바 그 결과는 다음과 같다. 1. 전품종의 평균 감소율은 질산환원효소의 활성이 42%이였으며 단백질분해효소의 활성은 73%였으며, 이에 반하여 proline은 대조구에 비하여 단수구가 무려 10배나 증가하였다. 2. 질산환원효소의 활성감소율을 맥종별로 보면 소맥 < 호맥 < 대맥 < 2조대맥ㆍ과맥의 순으로 소맥이 가장 낮았다. 3. 단백질분해효소의 활성감소율을 맥종별로 보면 소맥 < 호맥 < 이조대맥 < 대맥 < 과맥의 순으로 소맥이 높고 과맥이 가장 낮았다. 4. 단수구의 proline의 축적색대량을 맥종별로 보면 소맥 < 대맥 < 호맥 < 과맥 < 이조대맥의 순으로 소맥과 대맥이 높았으며, 대조구에 대한 단수구의 증가비는 호맥(13배) > 소맥ㆍ대맥(11배) > 과맥(9배) > 이조대맥(7배)의 순으로 호맥이 가장 높았다. 5. 한발저항성 과정에서 효소적 및 생리적 대사작용의 관점에서 볼 때 소맥 > 호맥 > 대맥 > 과맥 > 이조대맥의 순으로 한발저항성이 강한 것으로 추정된다.

• Current Research on Agriculture and Life Sciences
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• 제32권3호
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• pp.149-154
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• 2014

1. Pepsin과 pancreatin 소화물들을 비교한 결과, 순수한 팥 단백질 중에서 소화효소 저항성을 가지는 단백질이 존재하는 것으로 확인되었다. 2. 팥의 주요 단백질을 제거하고 pepsin과 pancreatin으로 소화시켰을 때 더 많은 분해가 일어나는 것으로 보아 팥의 주요단백질들 중에 소화효소 저항성을 가지는 것이 많을 것이라 추측된다. 3. 팥의 주요 단백질들은 장 점막세포와는 크게 작용하지 않는 것을 확인할 수 있었다. 4. 팥 단백질의 데이터베이스 구축과 팥 단백질이 다른 영양소들과의 상호작용을 하는 지에 대한 연구가 진행되어야 할 것이다.

• The Plant Pathology Journal
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• 제21권4호
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• pp.402-405
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• 2005

We have previously identified genes for four different protease inhibitors (PIs) that were induced upon rice blast infection in a rice blast resistant mutant SHM-11. Our expression analysis of the PIs indicated that induction of the PIs was the highest 24 hr after rice blast inoculation in the rice mutant SHM-11. Three PIs in the group of serine PIs were highly expressed while a cystein PI was weakly expressed upon rice blast inoculation. Four PIs were weakly induced 48 hr after pathogen inoculation in rice blast susceptible wild type rice plant. The simultaneous expression of three serine PIs was apparent from SHM-11 and two of them were induced in rice blast resistant Taebaegbyeo. One of them was induced in rice blast resistant Hwayeongbyeo while none of them were expressed in rice blast susceptible Nagdongbyeo and rice blast resistant Dongjinbyeo. Our results suggest that the expression of PI gene is rice cultivar specific and may be linked with the rice blast resistance in a specific rice mutant by the simultaneous expression of the PI genes.

• Journal of Microbiology
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• 제33권4호
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• pp.344-349
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• 1995

The alkalophilic bacterium, Xanthomonas sp. JK311, producing extracellular proteases, was isolated from soil. Xanthomonas sp. JK311 produced five extracellular proteases that are all metalloproteases. Four of them were resistant against 1% SDS. Chromosomal DNA of the Xanthomonas sp. JK311 was digested with BamHI and cloned into PUC19. Among E. coli strain HB101 transformants, a clone secreting the proteases was screened through halo formation on skim-milk agar plate and by Southern blot analysis. It had the recombinant plasmid pXEP-1 containing the 7.5 kb-BamHI DNA fragment and produced three extacellular proteases. Their protease properties corresponded to those of Xanthomonas sp. JK311.

• 한국미생물·생명공학회지
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• 제7권3호
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• pp.157-164
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• 1979

Aspergillus 속균중 Asp. awamori U-3가 acid protease의 생산능이 우수함으로 효소생산에 미치는 배양조건 및 조효소의 특성을 검토하고 다음과 같은 결과를 얻었다. 1. 최적배양온도는 3$0^{\circ}C$, 최적배양시간은 72 시간, 최적첨수량은 wheat bran medium 에서는 100~120 %, defatted rice bran medium 에서는 100~130%이었다. 2. 명종 성분중 wheat bran medium에는 KN $O_3$, glutamic acid, glucose 가, defatted rice bran me-dium 에는 KN $O_3$, (N $H_4$)$_2$S $O_4$, glucose, lactose, K $H_2$P $O_4$ 및 MgC $l_2$의 첨가가 특히 효과적이었으며, wheat bran medium 에서 가장 효과적인 glucose, KN $O_3$와 glutamic acid의 최적 첨가농도는 명명 3.0~4.0%, 0.2~0.4%, 1.0 %이었다. 3. 본효소의 작용 최적 pH는 2.4, 최적온도는 45$^{\circ}C$내외, 안정 pH 범위는 2.0~5.0이고, 5$0^{\circ}C$ 이하에서는안정하나 그 이상에서는 급격하게 불활성화 되었다. 4. 본효소에 대한 명종 무기염류의 내열효과를 검토한바 CaC $l_2$와 CaS $O_4$ 가 약간의 내열효과를 나타내었다. 5. 내열제를 첨가하고 6$0^{\circ}C$에서 10~30 분간 처리할때 20분 이상에서는 효소의 잔존활성이 크게 감소되었고, 내열제로서 CaC $l_2$와 CaS $O_4$ 를 첨가할 때 다같이 8$0^{\circ}C$에서는 내열효과는 거의 없었다.

• 한국식물생명공학회:학술대회논문집
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• 한국식물생명공학회 2005년도 추계학술대회 및 한일 식물생명공학 심포지엄
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• pp.219-219
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• 2005

• 대한바이러스학회지
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• 제28권4호
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• pp.295-302
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• 1998

The RNA genome of poliovirus encodes a long polyprotein precursor and this polyprotein is cleaved proteolytically by viral protease to yield mature proteins. The mature proteins derived from the P1 polyprotein precursor are the component of capsids. To further delineate the process of capsid assembly and encapsidation, in a first attempt, a cell line which expresses the authentic P1 polyprotein was established. CV-1 cells were transfected with the pRCRSVS1P1 plasmid DNA which contains 5'ncr sequences, whole authentic capsid gene of poliovirus and neomycin resistance gene. These cells were treated with G418 for 3 months, and eventually G418 resistant cells were selected and formed colonies. Each colony was picked and grown in the media containing G418. DNA analysis indicated that 1 of 13 neomycin resistant cell lines (R2-18) contains whole poliovirus P1 capsid gene segment which was incorporated into the genome. Immuneprecipitation of cell lysates with sera from rabbit immunized with inactivateded Sabin type 1 particles demonstrated the constitutive expression of the poliovirus P1 capsid protein from R2-18.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2003년도 생물공학의 동향(XII)
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• pp.180-184
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• 2003

선별된 7종의 균주는 넓은 범위의 산과 담즙산에 대해 기존에 보고된 결과에 비하여 높은 내성을 나타냈고, 장내세균인 E. coli에 대해서도 우수한 항균력을 나타냈고, 항생제 tetracycline, nisin, kanamycin, ampicillin. streptomycin에 대해서도 높은 내성을 보였다. 또한 7종의 균주는 어느 정도의 산을 생성하고 있었고, 네 가지 효소 amylase, protease, lipase, cellulase에 대해서도 높은 활성을 나타냈다. 균주 특성실험 결과, 7종의 균주는 그람음성균으로 모두 구형, 비운동성이고 균체의 색은 노란색이거나 흰색을 띠고 있고, 다양한 유기물에 대하여 분해력을 지니고 있었다.

• Asian-Australasian Journal of Animal Sciences
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• 제16권12호
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• pp.1822-1829
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• 2003

Porcine colostrum and milk were separated into the acid-soluble and casein fractions by acidification followed by centrifuge. The acid-soluble fraction of porcine colostrum was further separated by liquid chromatography and anisotropic membrane filtration. Trypsin and chymotrypsin inhibitory capacity in porcine colostrum, milk and their components was determined by incubating bovine trypsin or chymotrypsin in a medium containing their corresponding substrates with or without addition of various amounts of porcine colostrum, porcine milk or their components. The inhibition of insulin-like growth factor I (IGF-I) and epidermal growth factor (EGF) degradation in pig small intestinal contents by porcine colostrum was measured by incubating iodinated IGF-I or EGF with the intestinal contents with or without addition of porcine colostrum. Degradation of labeled IGF-I or EGF was determined by monitoring the generation of radioactivity soluble in 30% trichloroacetic acid (TCA). The results showed that porcine colostrum had high levels of trypsin and chymotrypsin inhibitory activity and increased the stability of IGF-I and EGF in pig intestinal contents. The inhibitory activity declined rapidly during lactation. It was also found that trypsin and chymotrypsin inhibitory activity and the inhibition on IGF-I and EGF degradation in the acid-soluble fraction were higher than that in the casein fraction. Heat-resistance study indicated that trypsin inhibitors in porcine colostrum survived heat treatments of $100^{\circ}C$ water bath for up to 10 min, but exposure to boiling water bath for 30 min significantly decreased the inhibitory activity. Compared with the trypsin inhibitors, the chymotrypsin inhibitors were more heatsensitive. Separation of the acid-soluble fraction of porcine colostrum by liquid chromatography and anisotropic membrane filtration revealed that the trypsin and chymotrypsin inhibitory capacity was mainly due to a group of small proteins with molecular weight of 10,000-50,000. In conclusion, the present study confirmed the existence of high levels of protease inhibitors in porcine colostrum, and the inhibition of porcine colostrum on degradation of milk-borne growth factors in the pig small intestinal tract was demonstrated for the first time.