• 공업화학
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• 제18권1호
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• pp.10-16
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• 2007

본 연구에서는 인체의 해로운 요소를 분해하고 분해 생성물인 암모니아($NH_3$)의 제거를 위한 우레아제 고정막 제조와 가수분해 특성에 관하여 검토되었다. 우레아제 고정막은 방사선 그라프트 중합법에 의해 다공성 중공사막에 음이온 교환기로서 DEA를 도입한 후 요소의 가수분해를 위해 우레아제를 고정시켜 제조하였다. 음이온 교환기가 도입된 막에 우레아제가 고정된 경우 그라프트율이 증가할수록 고정량이 증가하였다. 이것은 그라프트율이 증가할수록 그라프트 체인이 신장하여 단백질이 다층으로 고정되기 때문이다. 한편, 가교제의 첨가는 우레아제 고정막의 세척 과정에서 발생하는 탈리 현상을 억제할뿐 아니라 막의 반복사용도 가능하게 하였다. 5 h의 가교 반응을 통하여 탈리현상이 발생되지 않는 우레아제 고정막을 제조하였으며, 이때 제조된 우레아제 고정막은 1 mol과 4 mol의 요소 용액에서 각각 98%와 50% 이상의 가수분해 성능을 나타내었다.

• 한국막학회:학술대회논문집
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• 한국막학회 2004년도 추계 총회 및 학술발표회
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• pp.43-45
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• 2004

• Journal of Microbiology and Biotechnology
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• 제11권2호
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• pp.317-325
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• 2001

For heuristic purposes, the relative ratio of urease contents inside and outside cells was surveyed using nine ureB+ strains of Helicobacter pylori. the ratio of the enzyme specific activity appeared to vary greatly between the various H. pylori strains, ranging from 0.5 to 2.5. Besides the above compartment, urease was also richly found in the membrane fraction, especially in either peripheral or integral form. The urease distribution across the H. pylori membrane was significantly influenced by the ambient pH; the specific activity of external urease was highest at pH 5.5 with a narrow plateau, whereas the internal specific activity was highest within a pH range of 4.5 to 6.5 with a broad plateau. These finding strongly suggest that H. pylori urease is secretory and responded to the external pH. However, at pH 4.0 or below, no urease activity was detected in either the internal or external compartment, although an increase in the color development with 2,4,6-trinitrobenzene sulfonate (TNBS) was observed. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) demonstrated that these phenomena may be related to a specific proteolysis in certain proteins, including urease or ${\gamma}$-glutamyl transpeptidase. Interestingly, the effect of ammonium ions n alleviating the enzyme inactivation inside the H. pylori cells was remarkably similar to that of D-glucose. In addition, it would appear that the cation acted as a surrogate of not only $Na^+$ but also $K^+$ thereby increasing the H. pylori P-type ATPase activity. This is of great interest, as it implies that the urease action in H. pylori is indispensible at any locus.

• 한국막학회:학술대회논문집
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• 한국막학회 2004년도 춘계 총회 및 학술발표회
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• pp.49-52
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• 2004

생체 내에서의 요소 형성은 단백질이 아미노산으로 분해되어 인체에 남은 요소는 오줌으로 배출된다. 그러나 고농도의 urea의 경우 단백질을 변형시키게 된다[1-2]. 이러한 고 농도의 urea를 단백질 공정을 통해서 제거시키는 기술이 최근의 투석 과학이다. 그러나 이러한 방법은 urea의 제거와 함께 많은 양의 단백질과 양이온이 유출 및 오염의 문제가 많이 발생하고 있다[3].(중략)

• 생명과학회지
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• 제18권2호
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• pp.241-248
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• 2008

축산물을 이용하여 생산된 특이항체는 세균성감염에 의한 설사병 치료와 충치예방에 효과가 있고, 특히 계란을 이용한 IgY (Immunoglobulin Yolk)는 비교적 산과 열에 안정하다는 실험결과가 보고되어[22] 있다. 식품분야에 특이항체를 식품소재로 산업화에 활용한 빈도는 아직 미미한 상태에 있으나, 최근 면역학, 단백질공학, 생명공학 등의 발전과 기술 향상으로 식품 소제로써의 활용성이 점차 높아질 것으로 기대된다. 본 연구에서는 H. pylori의 감염을 예방하고 치료보조제로 사용할 목적으로 포유동물을 통한 피동면역용 항체를 생산하고자 하였다. H. pylori로부터 항원을 분리하고, 분리된 항원에 대한 항체생산 및 H. pylori의 응집정도를 알아보았다. H. pylori로부터 분리된 주요 항원 단백질은 WC, OMP, crude urease, LPS 각각 12개, 7개, 3개, 1개 종류의 band를 확인할 수 있었다. 분리된 항원의 IgG 항체 생성은 동일한 항원농도인 $20{\mu}g/100{\mu}l$에서 각각 WC (L) $77.9{\pm}6.4{\mu}g/ml$, OMP $84.9{\pm}6.4{\mu}g/ml$, crude urease $123.8{\pm}2.9{\mu}g/ml$로 crude urease 항원이 가장 많은 것으로 나타났다. 그리고 IgA 항체 생성은 WC (L) $2.5{\pm}0.32{\mu}g/ml$, OMP $2.0{\pm}0.43{\mu}g/ml$, crude urease $1.3{\pm}0.25{\mu}g/ml$로 IgA 항체 생성은 WC (L) 항원이 가장 많은 것으로 나타났다 Western blotting을 통하여 항원 단백질의 면역원성 알아본 결과 WC 10종류, OMP 6종류, crude urease 3종류의 주요 항원성 물질을 확인할 수 있었다. 항체의 H. pylori에 대한 응집정도를 나타내는 응집가는 anti-WC (H), anti-WC (L), anti-OMP, anti-crude urease 항혈청에서 각각 $2^5,\;2^5,\;2^6\;and\;2^7$으로 나타났으며, 상대적으로 anti-crude urease 항혈청이 가장 높은 응집가를 나타내었다. 각 항원에 의해 생성된 항혈청의 urease활성 억제에 대한 흡광도(OD=550 nm)는 WC (H) $0.14{\pm}0.01$, WC (L) $0.16{\pm}0.01$, OMP $0.18{\pm}0.03$, Urease $0.18{\pm}0.04$로 대조구 $0.26{\pm}0.02$와 비교할 때 유의적인 억제효과가 있는 것으로 나타났다. 이상의 결과를 종합해 볼 때 H. pylori 항원의 분리와 분리된 항원의 항체 생성능, H. pylori의 응집가, urease 활성억제측면에서 WC 및 crude urease항원 모두 높은 항체 역가를 나타내었다.

• Journal of Microbiology and Biotechnology
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• 제13권5호
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• pp.673-679
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• 2003

Glutamine and urea, abundant in body fluids or plasma, yield net ammonium ions upon hydrolysis by ${\gamma}-glutamyl$ transpeptidase (${\gamma}-GTP$) and urease, respectively, and these two enzymes are largely produced from Helicobacter pylori. To investigate bacterial potential of ammonium production, we first quantified those in whole-cell systems and found that the relative ratio of their amounts varied greatly, especially with pH values and the cell's aging. During the H. pylori cultivation, the ratio appeared to be inversely proportional to each other, showing a progressive increase of the ${\gamma}-GTP$ with decreasing of the urease. Under the urease-defective conditions due to low pH or coccoids, the bacterial cells still possessed a considerable amount of ${\gamma}-GTP$, which was found exclusively in the external compartment, therefore, the cell's ammonium production was found to be solely dependent upon glutamine, and the external ammonium concentration was constant without any contribution of urea concentration. Such ammonium constancy would definitely have an adverse effect on the host, because of its absolute requirement for vacuolar degeneration by H. pylori VacA, maximized at approximately 10 mM $NH_4Cl$. It was also found that, by using the metal-saturated membrane vesicles, ammonium ions were likely to be involved in the pH-dependent cation-flux across the H. pylori membrane, where the role of ${\gamma}-GTP$ in ammonium homeostasis around cells was suggested, especially under the hostile conditions against H. pylori.

• BMB Reports
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• 제30권5호
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• pp.308-314
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• 1997

We investigated biotinylation of nitrocellulose membrane for immuno-filtration capture assay. In order to enhance the efficiency of biotinylation, nitrocellulose membranes were pretreated with several chemicals for the purpose of suitable protein absorption through surface modification. As a signal generating enzyme, urease was used and the concentration of avidin was optimized for the efficient binding kinetics between urease-biotin in liquid phase and biotinylated membrane in solid phase. For effective biotinylation, bovine serum albumin-biotin complexes could be immobilized at a concentration of $370\;{\mu}g$/stick ($4.4\;cm^2$). Among tested chemicals, polylysine (0.25%) showed a significant effect in biotinylation. Polylysine is thought to enhance surface area by extending unbound residues into solution. Time of treatment over 30 min and higher molecular weight of polylysines (58,100 dalton) showed positive effect on the enhancement of biotinylation. The result from this study may be useful for developing a new biosensor and other biofunctional membranes for examining molecular recognition.

• 한국미생물·생명공학회지
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• 제25권2호
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• pp.129-136
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• 1997

Helicobacter pylori is a spiral-shaped, microaerophilic human gastric pathogen causing chronic-active gastritis in association with duodenal ulcer and gastric cancer. To investigate the possibility of H. pylori outer membrane proteins (OMPS) as the oral vaccine antigens, sarcosine-insoluble outer membrane fraction has been prepared from H. pylori NCTC 11637. The major OMPs having apparent molecular masses of 62 kDa, 54 kDa and 33 kDa were detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), which were identified as urease B subunit (UreB), heat shock protein (Hsp54 kDa) and urease A subunit (UreA), respectively. Minor protein bands of 57 kDa, 52 kDa, 40 kDa, 36 kDa and 31 kDa were also observed. The antigenicity of H. pylori OMPs and antigenic cross-reactivity among the strains were determined by immunoblot analysis using anti-H. pylori OMPs antisera or intestinal lavage solutions. The results showed that UreB, Hsp54 kDa, UreA and 40 kDa proteins vigorously stimulated mucosal immune response rather than systemic immunity. From this results, these proteins seemed to be useful as the antigen candidates for the oral vaccine. The immunoblotting results with surface proteins from eight isolated H. pylori strains were similar to that of H. pylori NCTC 11637. The IgA which had been arised from oral administration of H. pylori OMPs, was able to bind H. pylori whole-cells.

• Childhood Kidney Diseases
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• 제25권2호
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• pp.112-116
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• 2021

Hyperammonemia is mainly caused by diseases related to liver failure. However, there are also non-hepatic causes of hyperammonemia, such as urinary tract infection (UTI) due to urease-producing organisms. Urease production by these bacteria induces a hydrolysis of urinary urea into ammonia that can cross the urothelial cell membrane and diffuse into blood vessels, leading to hyperammonemia. Delayed diagnosis and treatment of hyperammonemia can lead to lethal encephalopathy that can cause brain damage and life-threatening conditions. In the presence of obstructive uropathy, UTI by urease-producing bacteria can lead to more severe hyperammonemia due to enhanced resorption of ammonia into the systemic circulation. In this report, we present a case of acute severe hyperammonemic encephalopathy leading to brain death due to accumulation of ammonia in blood caused by Morganella morganii UTI in a 10-year-old girl with cloacal anomaly, causing obstructive uropathy even after multiple corrections.

• 대한화학회지
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• 제36권3호
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• pp.393-404
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• 1992

새로운 효소반응기를 쓰는 요소 정량용 연속${\cdot}$자동화 장치의 감응성질을 조사하였다. 효소반응기는 지지체인 nylon-6입자(42∼48 mesh)를 teflon관(안지름 2mm, 길이 20cm)에 충전시키고, 이 지지체의 표면에 공유결합제인 glutaraldehyde로 urease를 고정화시켜서 제작하였다. 연속${\cdot}$자동화장치는 효소반응기, 기체투석기 및 지시전극인 관형 PVC-nonactin막 암모늄 이온 선택성 전극을 차례로 연결하여 만들었다. 이 장치를 써서 요소를 정량할 때 감응특성은 다음과 같다. 곧 직선감응 농도범위는 $5.5{\times}10^{-6}$~$2.4{\times}10^{-3}M$, 감응기울기는 57.8 mV/decade, 검출한계는 $1.5{\times}10^{-6}M$, 효소반응기의 전환백분율은 80.8%이었다. 효소반응기의 최적 완충용액은 0.01M Tris-HCl 완충용액(pH 7.0∼7.8)과 0.01M 인산염 완충용액(pH 6.9∼7.5)이었고, 수명은 150일 정도였다. 또한 다른 생리활성물질의 방해는 없었다.