• Reproductive and Developmental Biology
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• 제38권2호
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• pp.63-70
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• 2014

Plasminogen activators (PAs) are serine proteases that convert plasminogen to plasmin. Two type of PAs are urokinase-type PA (uPA) and tissue-type PA (tPA). Plasminogen is present in most extracellular fluids. PAs play in various reproductive processes including implantation, ovulation and fertilization. In the spermatozoa, PAs and PAIs play a role in sperm motility and fertilization. PAs in the sertoli cell are stimulated spermatozoa maturation and sperm activation through the phospholipase A2. The oocyte maturation is the process for fertilization and implantation. PAs in cumulus-oocyte complexes (COCs) are related to oocyte maturation by protein kinase A and C. In the ovulatory process, PAs activity are changed and it are related to reducing the tensile strength of ovarian follicle wall. The uterine environment is important for reproduction and the uterus undergo tissue remodeling. In the uterus and oviduct of mammals, expression and activity of PAs are changed during estrous cycle. Thus, expression and activity of PAs are concerned to many reproductive functions. Therefore, PAs seem to important factor of regulator in reproductive events.

• The Korean Journal of Physiology and Pharmacology
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• 제18권1호
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• pp.73-78
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• 2014

Cell death and survival are tightly controlled through the highly coordinated activation/inhibition of diverse signal transduction pathways to insure normal development and physiology. Imbalance between cell death and survival often leads to autoimmune diseases and cancer. Death receptors sense extracellular signals to induce caspase-mediated apoptosis. Acting upstream of CED-3 family proteases, such as caspase-3, Bcl-2 prevents apoptosis. Using short hairpin RNAs (shRNAs), we suppressed Bcl-2 expression in Jurkat T cells, and this increased TCR-triggered AICD and enhanced TNFR gene expression. Also, knockdown of Bcl-2 in Jurkat T cells suppressed the gene expression of FLIP, TNF receptor-associated factors 3 (TRAF3) and TRAF4. Furthermore, suppressed Bcl-2 expression increased caspase-3 and diminished nuclear factor kappa B (NF-${\kappa}B$) translocation.

• Asian-Australasian Journal of Animal Sciences
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• 제25권6호
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• pp.852-860
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• 2012

An Antarctic bacterial isolate displaying extracellular ${\alpha}$-galactosidic activity was named Paenibacillus sp. LX-20 based on 16S rRNA gene sequence analysis. Optimal activity for the LX-20 ${\alpha}$-galactosidase occurred at pH 6.0-6.5 and $45^{\circ}C$. The enzyme immobilized on the smart polymer Eudragit L-100 retained 70% of its original activity after incubation for 30 min at $50^{\circ}C$, while the free enzyme retained 58% of activity. The enzyme had relatively high specificity for ${\alpha}$-D-galactosides such as p-nitrophenyl-${\alpha}$-galactopyranoside, melibiose, raffinose and stachyose, and was resistant to some proteases such as trypsin, pancreatin and pronase. Enzyme activity was almost completely inhibited by $Ag^+$, $Hg^{2+}$, $Cu^{2+}$, and sodium dodecyl sulfate, but activity was not affected by ${\beta}$-mercaptoethanol or EDTA. LX-20 ${\alpha}$-galactosidase may be potentially useful as an additive for soybean processing in the feed industry.

• Biomolecules & Therapeutics
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• 제20권2호
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• pp.133-143
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• 2012

Matrix metalloproteinases (MMPs) are a subfamily of zinc-dependent proteases that are re-sponsible for degradation and remodeling of extracellular matrix proteins. The activity of MMPs is tightly regulated at several levels including cleavage of prodomain, allosteric activation, com-partmentalization and complex formation with tissue inhibitor of metalloproteinases (TIMPs). In the central nervous system (CNS), MMPs play a wide variety of roles ranging from brain devel-opment, synaptic plasticity and repair after injury to the pathogenesis of various brain disorders. Following general discussion on the domain structure and the regulation of activity of MMPs, we emphasize their implication in various brain disorder conditions such as Alzheimer's disease, multiple sclerosis, ischemia/reperfusion and Parkinson's disease. We further highlight accumu-lating evidence that MMPs might be the culprit in Parkinson's disease (PD). Among them, MMP-3 appears to be involved in a range of pathogenesis processes in PD including neuroinflamma-tion, apoptosis and degradation of ${\alpha}$-synuclein and DJ-1. MMP inhibitors could represent poten-tial novel therapeutic strategies for treatments of neurodegenerative diseases.

• Parasites, Hosts and Diseases
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• 제41권4호
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• pp.189-196
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• 2003

In order to evaluate the possible roles of secretory proteases in the pathogenesis of amoebic keratitis, we purified and characterized a serine protease secreted by Acanthamoeba lugdunensis KA/E2, isolated from a Korean keratitis patient The ammonium sulfate-precipitated culture supernatant of the isolate was purified by sequential chromatography on CM-Sepharose, Sephacryl S-200, and mono Q-anion exchange column. The purified 33 kDa protease had a pH optimum of 8.5 and a temperature optimum of $55^{\circ}C$. Phenylmethylsulfonylfluoride and 4-(2-Aminoethyl)-benzenesulfonyl-fluoride, both serine protease specific inhibitors, inhibited almost completely the activity of the 33 kDa protease whereas other classes of inhibitors did not affect its activity. The 33 kDa enzyme degraded various extracellular matrix proteins and serum proteins. Our results strongly suggest that the 33 kDa serine protease secreted from this keratopathogenic Acanthamoeba play important roles in the pathogenesis of amoebic keratitis, such as in corneal tissue invasion, immune evasion and nutrient uptake.

• 한국미생물·생명공학회지
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• 제51권2호
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• pp.167-173
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• 2023

Proteolytic enzymes secreted by Aspergillus, as pathogenicity factors, affect blood coagulation and fibrinolysis, and therefore the target proteins of their action in the bloodstream are of significant interest. In the present study, the action of the isolated protease of A. terreus 2 on different human plasma proteins was shown. The protease of A. terreus 2 exhibited the highest proteolytic activity against hemoglobin, which was 2.5 times higher than the albuminolytic activity shown in both of the protein substrates used. In addition, the protease has significant ability to hydrolyze both fibrin and fibrinogen. However, the inability of the A. terreus 2 protease to coagulate rabbit blood plasma and coagulate human and bovine fibrinogen indicates the severity of the enzyme's action on human blood coagulation factors. It should be considered as a potential indicator of this isolated protease's participation in fungal pathogenesis. The protease shows no hemolytic activity. Furthermore, its activity is insignificantly inhibited by thrombin inhibitors, and is not inhibited by plasmin inhibitors.

• International Journal of Industrial Entomology and Biomaterials
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• 제22권2호
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• pp.83-93
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• 2011

Biochemical and enzymatic characterization for extracellular protease isolated from Cordyceps militaris cultivated on rice bran medium was investigated. C militaris produced proteolytic enzymes from 10 days after inoculation, maximum enzyme production was found at 25 days. The optimum temperature and pH of proteases production was at $25^{\circ}C$ and pH 7.0, respectively. The protease activity was observed in the four peaks (Pro-I, Pro-II, Pro-III, and Pro-IV) separated through Sephadex G-100 column chromatography. The separated protease was optimally active at $25^{\circ}C$. Optimum pH of the protease was between 7 and 8. Enzyme was also stable over at $30-80^{\circ}C$. The enzyme was highly stable in a pH range of 4-9. Protease activity was found to be slightly decreased by the addition of $Mg^{2+}$, $Mn^{2+}$, $Zn^{2+}$, $Fe^{2+}$ and $Cu^{2+}$, whereas inhibited by the addition of $Ca^{2+}$ and $Co^{2+}$ Protease activity was inhibited by protease inhibitor PMSF. On the other hand, the partially purified protease was investigated on proteolytic protease activity by zymogram gel electrophoresis using three substances (casein, gelatin and fibrin). Four active bands (F-I, FII, F-III, and F-IV) of fibrin degradation were revealed on fibrin zymogram gels. Both of F-II and FIII showed caseinolytic, fibrinolytic and gelatinolytic activities in three gels. Thermostability, pH stability, and pH-thermostability of the enzyme determined the residual fibrinolytic activity also displayed on fibrin zymogram gel. The only one enzyme (F-II) displayed over a broad range of temperature at $30-90^{\circ}C$. The FII displayed fibrinolytic activity in the pH range 3-5, but was inactivated in the range of pH 6-11. The F-I and F-III showed enzyme activity in the pH range of 6-11. In the pH-thermostability, the F-II only kept fibrinolytic activity after heating at $100^{\circ}C$ for 10, 20 and 30 min at pH 3 and pH 7, respectively. On the other hand, the F-II was retained activity until heating for 10 min under pH 11 condition. By using fibrin zymogram gel electrophoresis, extracellular fibrinolytic enzyme F-II from C. militaris showed unusual thermostable under acid and neutral conditions.

• 대한화장품학회지
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• 제33권4호
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• pp.245-249
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• 2007

노화 과정 중에 일어나는 extracellular matrix (ECM)의 변성은 피부의 주름과 탄력 감소를 유발한다. 현재까지 항노화의 주요 타겟은 metalloproteases 혹은 콜라겐이나 엘라스틴 같은 구조 단백질에 집중되어 있지만, 최근 세포와 ECM 단백질(콜라겐, 피브릴린, 파이브로넥틴) 간의 상호작용이 세포의 생존과 증식, 조직의 재건에 중요한 역할을 한다고 알려졌다. 파이브로넥틴은 다른 ECM 단백질이나 인테그린 같은 세포 표면 수용체와 결합할 수 있는 부위를 가진 부착 단백질이다. 최근 보고에 따르면 세린 프로티아제들에 의해 분해된 파이프로넥틴 조각이 골아세포에서 MMPs 발현을 증가시킨다. 그러나 파이브로넥틴 조각의 사람 피부에서의 역할은 보고된 바 없다. 본 연구에서는 노인의 피부에서 파이브로넥틴 조각이 현저히 증가되어 있으며, 섬유아세포에 파이브로넥틴 조각을 처리하였을 시, MMP-1의 발현과 MMP-2의 활성이 증가한다는 것을 입증하였다. 이 결과는 파이브로넥틴 조각이 피부 노화를 유발하는 새로운 인자일 가능성을 제시하고 있다.

• 생명과학회지
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• 제25권2호
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• pp.214-222
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• 2015

Aeromonas hydrophila는 수생계에 흔히 존재하는 병원체로서 출혈성 패혈증, 수종, 궤양 등의 질병을 유발한다. A. hydrophila는 효과적인 감염과 다양한 환경에서 적응하기 위해 세포 외 물질(Extracellular products, ECPs)을 분비한다. 본 연구에서 사용한 A. hydrophila CK257은 매우 독성이 강한 균주로 균이 분비하는 ECP 역시 쥐, 토끼, 금붕어를 포함한 동물 모델에게 조직 손상을 일으키며 독성을 보였다. 이러한 괴사 반응을 일으키는 원인 요소를 찾기 위해 ECP의 특성을 분석하였고 그 결과, 원인 요소가 단백질이라는 것을 확인하여 이후 단백질 분리 및 정제에 초점을 맞추어 실험을 진행하였다. 균체를 제거한 배양액 상의 분비 단백질을 얻기 위하여 ammonium sulfate 침전법을 사용하였고, 이후 겔 거르기 크로마토그래피(Gel filtration chromatography)를 이용하여 단백질을 정제하였다. 정제 후 단백질은 다섯 개의 분획으로 나뉘었고 이 중 한 개의 분획이 동물 모델에 괴사와 같은 피부 손상을 일으키는 것을 확인하였다. 괴사 활성을 가진 분획의 단백질 4개의 밴드를 MALDI-TOF 시스템을 통해 분석한 결과, Peptidase M35와 Peptidase M28로 밝혀졌으며 이들은 각각 금속촉매효소(metalloprotease)임을 확인할 수 있었다. Peptidase M35와 Peptidase M28의 단백질 분해 효소로써의 기능을 확인하기 위해 카제인 분해 활성을 확인하였고, 다양한 금속 이온을 처리함에 따라 그 활성이 달라지는 것을 관찰할 수 있었다. 또한 조직에 존재하는 단백질의 한 종류인 엘라스틴(elastin) 분해능 역시 확인해 보았다. 본 연구에서는 A. hydrophila가 분비하는 물질 가운데 조직 괴사 활성을 일으키는 것으로 추정되는 Peptidase M35와 Peptidase M28의 두 가지 인자를 확인하여 동정하였다. 이후 두 가지 인자를 결손 시킨 돌연변이주를 구축하여 조직 괴사에 이들이 직접적으로 작용하는지 확인 할 예정이다.

• BMB Reports
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• 제30권1호
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• pp.46-54
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• 1997

Alkaliphilic Bacillus sp. S-1 secretes a large amount (approximately 80% of total pullulanase activity) of an extracellular pullulanase (PUL-E). The pullulanase exists in two forms: a precursor form (PUL-I: $M_r$ 180,000), and a processed form (PUL-E: $M_r$ 140,000). Two forms were purified to homogeneity and their properties were compared. PUL-I was different in molecular weight, isoelectric point, $NH_2$-terminal amino acid sequence, and stabilities over pH and temperature ranges. The catalytic activities of PUL-I were also distinguishable in the $K_m$ and $V_{max}$ values for various substrates, and in the specific activity for pullulan hydrolysis. PUL-E showed 10-fold higher specific activities than PUL-I. However. PUL-I is immunologically identical to PUL-E, suggesting that PUL-I is initially synthesized and proteolytically processed to the mature form of PUL-E. Processing was inhibited by PMSF, but not by pepstatin, suggesting that some intracellular serine proteases could be responsible for processing of the PUL-I. PUL-I has a different conformational structure for antibody recognition from that of PUL-E. It is also postulated that the translocation of alkaline pullulanase(AP) in the bacterium possibly requires processing of the $NH_2$-terminal region of the AP protein. Processing of the precursor involves a conformational shift. resulting in a mature form. Therefore. precursor processing not only cleaves the signal peptide, but also induces conformational shift. allowing development of active form of the enzyme.