• Food Science and Biotechnology
• /
• 제15권1호
• /
• pp.28-32
• /
• 2006

Protease inhibitor of 72.6 kDa was successively purified from chum salmon (Oncorhynchus keta) eggs by ion exchange, gel permeation, and affinity chromatographies. Protease inhibitor was purified with yield and purification fold of 1.50% and 58.11, respectively. SDS-PAGE results showed purified protease inhibitor consisted of two protein subunits of 54.0 and 18.6 kDa. Chum salmon inhibitor exhibited stability between 20 and $40^{\circ}C$ in weak acid environment (PH 6), and inhibited papain and cathepsin, members of cysteine protease, but not chymotrypsin. The protein inhibited cathepsin more effectively than did egg white protease inhibitor, whereas the reverse was true for papain. These results indicate chum salmon egg inhibitor is heterodimer, thus the inhibitor was classified as cysteine protease inhibitor.

• Journal of Microbiology and Biotechnology
• /
• 제1권4호
• /
• pp.288-292
• /
• 1991

The aim of the current study is to evaluate the effects of medium compositions on the production of protease inhibitor in Streptomyces sp. SMF13. The production of protease inhibitor was counter-currently linked to extra-cellular protease, which were regulated by the culture conditions. Nitrogen source was the most critical ingredient affecting the production of protease inhibitor and protease. Carbon source was an important factor to determine the culture pH which affected very clearly the formation of protease and protease inhibitor. Inorganic phosphate inhibited the protease inhibitor production which was linked to the cell growth rate, although the optimal conditions for the production of protease inhibitor were not favouring to the cell growth.

• Journal of Microbiology and Biotechnology
• /
• 제16권4호
• /
• pp.524-530
• /
• 2006

Two protease inhibitors of 67 and 18 kDa, respectively, were purified from the eggs of glass fish, Liparis tanakai, by affinity chromatography and electro-elution method. The high molecular weight (HMW) protein was purified with a specific inhibitory activity, yield, and purity of 18.46 U/mg, 0.07%, and 131.86 fold, respectively, and was further characterized: Optimal temperature and pH for inhibitory activity of the HMW glassfish egg protease inhibitor were $40^{\circ}C$ and pH 6, respectively, and it was stable between $5^{\circ}C\;and\;50^{\circ}C$ in the pH range of 5-6 with maximal stability at pH 6. It was shown to be a competitive inhibitor against papain with an inhibition constant $(K_i)$ of 97.02 nM. Moreover, the 67 kDa protein inhibited cathepsin, a cysteine protease, more effectively than did an egg-white protease inhibitor. The HMW glassfish egg protease inhibitor was classified as a member of the family III (kininogen).

• Parasites, Hosts and Diseases
• /
• 제46권3호
• /
• pp.183-186
• /
• 2008

Helminthic cysteine proteases are well known to play critical roles in tissue invasion, nutrient uptake, and immune evasion of the parasites. In the same manner, the sparganum, the plerocercoid of Spirometra mansoni, is also known to secrete a large amount of cysteine proteases. However, cysteine protease inhibitors regulating the proteolytic activities of the cysteine protease are poorly illustrated. In this regard, we partially purified an endogenous cysteine protease inhibitor from spargana and characterized its biochemical properties. The cysteine protease inhibitor was purified by sequential chromatographies using Resource Q anion exchanger and Superdex 200 HR gel filtration from crude extracts of spargana. The molecular weight of the purified protein was estimated to be about 11 kD on SDS-PAGE. It was able to inhibit papain and 27 kDa cysteine protease of spargana with the ratio of 25.7% and 49.1%, respectively, while did not inhibit chymotrypsin. This finding suggests that the cysteine protease inhibitor of spargana may be involved in regulation of endogenous cysteine proteases of the parasite, rather than interact with cysteine proteases from their hosts.

• 한국미생물·생명공학회지
• /
• 제18권5호
• /
• pp.501-505
• /
• 1990

토양으로부터 분리한 Streptomyces 속 세균 KIS 13은 thiol 계통 단백질분해효소 활성을 특이적으로 저해하는 저분저량 저해물질을 생성하였다. 저해물질 생성은 세균체성장에 연관된 생성양상이 나타내었다. 배양액으로부토 butanol 추출, silicagel 60 column chromatography, Sephadex LH-2 gel-filtration chromatography, preparative HPLC 등의 과정을 통하여 단백질 분해효소 저해물질을 순수분리하였다. 이 저해물질은 Hammersten casein을 기질로 사용할때, papain에 대하여 non-competitive한 저해양상을 나타내었다.

• 미생물학회지
• /
• 제28권3호
• /
• pp.264-267
• /
• 1990

The objectives of the current studies were to establish the optimal conditions for the production of extracellular protease inhibitor in a strain of Streptomyces fradiae. As results, it was found that cell specific growth rate was very critical for the production of protease inhibitor and the optimum specific growth rate was found to be 0.05 h$^{-1}$ . Dissolved oxygen tension and pH were also important to regulate the inhibitor production. The inhibitory mode of the purified inhibitor to .alpha.-chymotrypsin was found to be competitive (K$_{i}$=5.5*10$^{-7}$ M). One mole of inhibitor could bind two moles of .alpha.-chymotrypsin and the complex has very low dissociation constant.t.

• 한국동물학회지
• /
• 제38권2호
• /
• pp.294-298
• /
• 1995

The genetic polymorphism of Protease inhibitor (Pl) in Korean population was investigated by using isoelectric focusing (IEF) in an ultra-narrow pH range,4.2-4.9, and immunoblottins. Three common alleles (Pl * Ml, Pl*2, Pl * M3) were observed and the frequencies for the alleles were Pl * M1=0.7843, Pl * M2=0.1613, Pl * M3=0.0323. In addition to the three common alleles, rare alleles (Pl *5, Pl * Z, Pl* H were detected at low-level frequency. Two unknovlm variants, which were not reported on previous studies in Korean population, were also found.

• 한국어병학회지
• /
• 제10권1호
• /
• pp.31-38
• /
• 1997

잉어로부터 분리한 Aeromonas hydrophila는 세포 밖으로 여러 종류의 proteases를 생산한다. A. hydrophila의 배양 상층액을 이용한 inhibitor assay를 통하여, 주된 활성을 나타내는 metallopretease와 약한 활성을 나타내는 serine protease가 있음을 알게 되었다. Gelatin SDS-PAGE를 통하여 두 개의 활성 band가 관찰되었으며, 이 들 중 넓게 퍼진 band는 metalloprotease에 특이하게 작용하는 inhibitor인 EDTA에 의해 활성이 상실되었고 따라서 metalloprotease임을 알 수 있었다. 다른 하나는 serine protease에 특이하게 반응하는 inhibitor인 PMSF에 의해 저해되어 serine protease임을 알 수 있었다. 이러한 두 extracellular protease의 활성은 $75^{\circ}C$에서 30 분간 열을 가한 후에도 Gelatin SDS-PAGE상에 남아있었다. 그런데, 주된 metalloprotease는 Sephadex G-75를 이용한 column chromatography를 거친 후 Gelatin Gel상에서 두 개의 band로 분리되었다.

• Applied Biological Chemistry
• /
• 제32권2호
• /
• pp.116-125
• /
• 1989

대두로부터 추출, 정제한 Bowman-Birk형 protease inhibitor 들의 함량과 전기영동양상을 비교하고 정제된 inhibitor내에 존재하는 각 isoinhibitor들의 함량을 조사하였으며 종실내 총 chymotrypsin 저해활성도와 cysteine 함량과의 관계를 알아보았다. 8품종의 대두로부터 Sephadex G-75를 이용하여 얻은 정제된 Bowman-Birk형 protease inhibitor들의 함량은 단백질 100g당 $6.67{\sim}9.36g$으로 품종간에 큰 차이가 있었다. 정제된 Bowman-Birk형 protease inhibitor들의 전기영동 양상은 품종간에 많은 차이를 보였으며 각 band의 함량에도 차이가 있었다. 8품종의 정제된 inhibitor에서 나타나는 총 14개의 전기영동 band 중 10개의 band가 protease 저해활성도를 보유하였고 그중 chymotrypsin 저해활성도가 높은 것은 band 7이었으며 그 함량은 단백질 100g당 $0.768{\sim}1.271g$으로 품종간에 차이를 보여 종실내 chymotrypsin 저해활성도가 높은 품종에서 함량이 많았다. 대두품종별 chymotrypsin 및 trypsin저해활성도는 각각 종실 g당 $52,200{\sim}15,225$와 $164,700{\sim}37,500$으로 품종간에 3배 및 4배의 차이를 보였고 단백질 g당으로 환산한 값은 $158,662{\sim}40,033$ 및 500, $608{\sim}120,347$로서 종실 g당 저해활성도와 같은 경향이었으며 3배 이상의 차이를 보였다. 품종별 cysteine 함량은 단백질 g당 $0.1258{\sim}0.0763-mmoles$로서 품종간에 차이를 나타내었으며 이는 실내 총 chymotrypsin 저해활성도와 정의상관관계가 있었다.

• Animal Bioscience
• /
• 제36권7호
• /
• pp.1034-1043
• /
• 2023

Objective: Two serine protease inhibitors, peptidase inhibitor 3 (PI3) and secretory leukocyte protease inhibitor (SLPI), play important roles in protease inhibition and antimicrobial activity, but their expression, regulation, and function at the maternal-fetal interface in pigs are not fully understood. Therefore, we determined the expression and regulation of PI3 and SLPI in the endometrium throughout the estrous cycle and at the maternal-fetal interface in pigs. Methods: Endometrial tissues during the estrous cycle and pregnancy, conceptus tissues during early pregnancy, and chorioallantoic tissues during mid to late pregnancy were obtained, and the expression of PI3 and SLPI was analyzed. The effects of the steroid hormones estradiol-17β (E2) and progesterone (P4) on the expression of PI3 and SLPI were determined in endometrial explant cultures. Results: PI3 and SLPI were expressed in the endometrium during the estrous cycle and pregnancy, with higher levels during mid to late pregnancy than during the estrous cycle and early pregnancy. Early-stage conceptuses and chorioallantoic tissues during mid to late pregnancy also expressed PI3 and SLPI. PI3 protein and SLPI mRNA were primarily localized to endometrial epithelia. In endometrial explant cultures, the expression of PI3 was induced by increasing doses of P4, and the expression of SLPI was induced by increasing doses of E2 and P4. Conclusion: These results suggest that the PI3 and SLPI expressed in the endometrium and conceptus tissues play an important role in antimicrobial activity for fetal protection against potential pathogens and in blocking protease actions to allow epitheliochorial placenta formation.