• 한국미생물·생명공학회지
• /
• 제27권6호
• /
• pp.458-465
• /
• 1999

Streptomyces luteogriseus KT-10 isolated from Korean farm soil produced a strong cathepsin B inhibitor. Optimal conditions for the cathepsin B inhibitor production by s. luteogriseus KT-10 were evaluated. The cathepsin B inhibitor was produced with maximal yield in the cultural condition of pH 7.0 and $25^{\circ}C$ for 4 days. Optimal medium for the cathepsin B inhibitor production was determined to be a medium containing 20g, peptone 3g, yeast extract 1g, K2HPO4 0.5g, MgSO4.7H2O 0.5g, NaNO3 0.5g, NaCl 0.5g per l. The cathepsin B inhibitor produced by S. luteogriseus KT-10 could also inhibit the other proteinases such as trypsin, papain, and cathepsin D.

• 한국미생물·생명공학회지
• /
• 제29권1호
• /
• pp.25-30
• /
• 2001

A strain of Actinomycetes producing cathepis B inhibitor was isolated from soil and identified as Streptomyces misakiensis. The product of S. misakiensis inhibited effectively cathepsis B proteinases as well as trypsin and papain. The cathepsin B inhibitor were largely produced with incubation for 4 days. The S. misakiensis was the most growth with incubation for 5 days. The cathepsin B inhibitor was isolated from the extraction of both with ethanol, ethanol and chlorofrom, and following several column chromatography such as sephadex G-15, silica gel 60 and sephadex LH-20 chromatography. The moleculer weight of purfied inhibitor was 138 dalton.

• 약학회지
• /
• 제39권3호
• /
• pp.297-305
• /
• 1995

The aim of the present study was to develop strains of actinomycetes producing low molecular weight cathepsin B inhibitor. Among 700 isolates from soil samples, a strain of Streptomyces sp. SMF30 producing cathepsin B inhibitor showing specificity and heat stability was selected by an economical and effective screening method. 50 units characteristics for major cluster analysis and 34 units characteristics for minor cluster were tested and the data were analyzed numerically using the TAXON program. The Isolate SMF30 was identified as a strain of Streptomyces aburabiensis

• 한국미생물·생명공학회지
• /
• 제29권2호
• /
• pp.84-89
• /
• 2001

포유동물조직의 세포내에 존재하는 lysosomal cysteine계 cathepsin B 효소는 암 발병과 전이, 류머티스 관절염, 염증 및 노인성치매 등의 여러 질병에 관여하고 있다. 이 cathepsin B를 저해하는 저해물질이 Streptomyces luteogriseus KT-10에 의해 생산되어 분리되었다. S. luteogriseus KT-10이 생산하는 cathepsin B 저해물질은 열에 안정하며 산, 알칼리에도 안정한 물질로 구성되어 있다. Cathepsin B 저해물질은 DEAE-Sephadex A-25, Sephadex G-15, silica gel, Sephadex LH-20의 column chromatography 과정을 거친 후 분취용 HPLC를 이용 분취한 후 백색분말의 cathepsin B 저해물질을 정제하였다. 이 정제한 저해물질은 UV 상의 전 범위에서 특이한 검출부위를 확인할 수 없었으며, $H_2$O, methanol, ethanol, n-butanol에는 녹지만 비극성인 chloroform, n-hexane, benzene 등에는 불용성이었다. 또한 ninhydrine, $H_2$$SO_4$, iodine에 양성 반응이지만 Ehrlichs, Pauly, Sakaguchi, phthalic acid, DNS, aniline 등의 단백반응과 당 반응에는 음성 반응으로 나타났다. IR 기기에서는 OH 기와 CH 기의 peak를 확인하였으며 $^1$H NMR 기기로 OH peak와 NH peak 또한 확인할 수 있었다. $^{13}$ C NMR spectrum은 4개의 탄소 peak를 가진 물질로 확인된 분자량이 207 dalton인 저해물질이다. 원소 분석기를 이용한 저해물질 분석 결과 분자식이 $C_4$$H_{11}$ $O_4$$N_{6}$로 나타났다. 이 cathepsin B 저해물질의 저해양식은 competitive 저해로 작용함을 확인할 수 있었다.

• 한국미생물·생명공학회지
• /
• 제23권5호
• /
• pp.602-608
• /
• 1995

The aim of the present research program was to construct an optimum fermentation system and to characterize the properties of cathepsin B inhibitor from Streptomyces chromofuscus SMF28. Glucose and casitone were proved to be good carbon source and nitrogen source, respectively. The production of inhibitor was high at lower concentration than 10 mM of inorganic phosphate. The optimum temperature and pH for the production of inhibitor were 30$\circ$C and pH 7, respectively. The production of inhibitor was related to mycelial growth and was affected by medium composition. The inhibitor in culture filtrate of S. chromofuscus SMF28 was purified by butanol extraction, silica gel chromatography, Amberlite IRC-50 (H$^{+}$ form) chromatography, preparative TLC, and preparative HPLC. From amino acid analysis and UV, IR, $^{1}$H-NMR spectroscopic analysis, the inhibitor was identified as a peptide containing valine and phenylalanine derivative.

• 한국미생물·생명공학회지
• /
• 제23권5호
• /
• pp.565-572
• /
• 1995

The aim of the present research program was to develop a strain of actinomycetes producing extracellular cathepsin B inhibitor. Soil samples were collected from various sites in Korea and a number of actinomycetes were isolated from the soil samples by applying various physical and chemical pretreatments. An economical and effective method was developed for the screening of strains producing low molecular weight cathepsin B inhibitor, and consequently a strain (SMF28) among over 700 isolates was selected. Chemotaxonomic and numerical identification were carried out for the isolate. Fifty taxonomic unit characters were tested and the data were analyzed numerically using TAXON program. The isolate was identified as a strain of Streptomyces chromofuscus.

• Journal of Microbiology and Biotechnology
• /
• 제18권6호
• /
• pp.1115-1120
• /
• 2008

A novel cathepsin B inhibitor-producing bacterium was isolated from marine sediments and identified based on its 16S rDNA sequence as Pseudomonas sp. strain PB01 (Accession No. EU126129). The growth and enzyme inhibitor production were investigated under various culture conditions. A mixture of organic nitrogen source was required for the optimal production, whereas both glucose and maltose proved to be the effective carbon sources for cathepsin B inhibitor production. Other optimal culture conditions included temperature range between 25 and $28^{\circ}C$, initial medium pH of 6.6, and shaking speed of 200 rpm. Under these optimal conditions, the maximum inhibitory activity from culture broth was approximately 50% after 30 h of cultivation. Additionally, kinetic study revealed that inhibitor production paralleled with cell growth, which suggested that the inhibitor may be a primary metabolite of that bacterium.

• 한국미생물·생명공학회지
• /
• 제29권2호
• /
• pp.90-95
• /
• 2001

Cysteine 계 cathepsin B 단백질 분해 효소는 고등동물의 세포조직 내에 lysosome에 존재하며 병원성세균의 침입을 막고 불필요한 단백질을 분해시키며 또한 면역세포의 항원인식에 대한 단백질 생산 등에 관여하는 효소이다. 이 cathepsin B의 과량 발현은 암전이, 만성적 염증성 질환, 류머티스 관절염, 노인성치매 등의 원인이 된다. 이 cathepsin B를 저해하는 것으로 밝혀진 S. luteogriseus KT-10으로부터 분리한 저해제 KHS10을 이용하여 분광광도계를 사용, 그 저해력을 조사하였다. Cathepsin B 저해제 KHS10은 경쟁적 저해를 나타내며 cathepsin B는 기질 CLN에 대해 Km 값은 0.5mM을 Vmax 값은 $29.4\mu$M로 나타내었으며, 합성기질인 BANA에 대해서는 Km 값이 2mM을 Vmax 값은 7.8$\mu$M/min로 나타났다. 저해제 KHS10의 Ki 값은 $0.43{\mu}$M로 측정되었다. Cathepsin B에 대한 저해제 KS10의 반응시간에 대한 저해력은 1시간 반응시 저해력이 100%에 가깝게 높게 나타났으며 온도에 의한 저해율은 $25^{\circ}C$에서 활성이 가장 높게 나타났다. Cathepsin B와 저해제 KHS10이 반응 전 배양은 5분간의 전 배양으로 높은 저해율을 나타내었으며 또한 pH에 의한 저해활성은 pH 6.0에서 가장 높게 저해하는 것으로 조사되었다. 저해제 KHS10는 $100^{\circ}C$에서 1시간 열을 가해도 그 잔류 저해력은 80% 이상 유지되었다.

• Restorative Dentistry and Endodontics
• /
• 제44권2호
• /
• pp.17.1-17.10
• /
• 2019

Objectives: Root resorption is an unexpected complication after replantation procedures. Combining anti-osteoclastic medicaments with retrograde root filling materials may avert this resorptive activity. The purpose of this study was to assess effects of a cathepsin K inhibitor with calcium silicate-based cements on osteoclastic activity. Methods: MC3T3-E1 cells were cultured for biocompatibility analyses. RAW 264.7 cells were cultured in the presence of the receptor activator of nuclear factor-kappa B and lipopolysaccharide, followed by treatment with Biodentine (BIOD) or ProRoot MTA with or without medicaments (Odanacatib [ODN], a cathepsin inhibitor and alendronate, a bisphosphonate). After drug treatment, the cell counting kit-8 assay and Alizarin red staining were performed to evaluate biocompatibility in MC3T3-E1 cells. Reverse-transcription polymerase chain reaction, tartrate-resistant acid phosphatase (TRAP) staining and enzyme-linked immunosorbent assays were performed in RAW 264.7 cells to determine the expression levels of inflammatory cytokines, interleukin $(IL)-1{\beta}$, IL-6, tumor necrosis $factor-{\alpha}$ ($TNF-{\alpha}$) and prostaglandin E2 (PGE2). Data were analyzed by one-way analysis of variance and Tukey's post hoc test (p < 0.05). Results: Biocompatibility results showed that there were no significant differences among any of the groups. RAW 264.7 cells treated with BIOD and ODN showed the lowest levels of $TNF-{\alpha}$ and PGE2. Treatments with BIOD + ODN were more potent suppressors of inflammatory cytokine expression (p < 0.05). Conclusion: The cathepsin K inhibitor with calcium silicate-based cement inhibits osteoclastic activity. This may have clinical application in preventing inflammatory root resorption in replanted teeth.

• Journal of Microbiology and Biotechnology
• /
• 제10권1호
• /
• pp.81-85
• /
• 2000

An elastase inhibitor, SMFEI02, was isolated from culture broth of Streptomyces lavendulae SMF11. The inhibitor was purified by ultrafiltration followed by XAD-7 column and Dowex-1 anion-exchange chromatographies, and preparative HPLC. The molecular formula was determined to be $C_{14}H_{16}N_2O_2$ (MW244) by HRFAB-MS analysis. The inhibitor was identified to be a diketopiperazine cyclo(S-Phe-S-Pro) by the optical rotation value and MNR spectral data, and showed inhibitory activities for trypsin, chymotrypsin, cathepsin B, and papain as well as elastase with the Ki values ranging from 1.78mM to $2.86{\;}\mu\textrm{m}$. The inhibition showed a competitive mode for elastase, chymotrypsin, and cathepsin B, whereas it showed a noncompetitive mode for trypsin and papain.