• 미생물학회지
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• 제30권6호
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• pp.546-552
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• 1992

에탄올 내성 및 고 알콜 생성 균주인 S. cerevisiae BU-1 와 glucoamylase 생성 우수 균주인 S. diastaticus A15 를 시험 균주로 하여 genetic manipulation 에 의하여 haploid 인 S. cerevisiae BUI.alpha.26($alc^r thr^-$) 와 S. diastaticus A15a6($STA^+ hom$) 균주를 분리하였다. 이등 효모 균주의 원형질체 형성을 위하여 0.8 M sorbitol 의 존재하에서 zymolyase $200\mu$g/ml 와 $400\mu$g/ml 을 2 시간 동안 처리시 두 균주 모두 95% 이상 원형질체를 얻었다. PEG 6000 을 90 분간 반응시킬 경우 융합 빈도는 $ 3.25 {\times} 10^{-3}$ 이었다. 원형질체 융합으로 생성된 융합주의 유전적 분석, 에탄올 내성, glucoamylase 생성을 측정한 결과, 이들 융합주의 유전자형은 STA $alc^r$ thr hom. '/STA' $alc^r$ thr hom로 예상되었으며, glucoamylase 활성도는 A15a6 의 경우 2.7 units 이었으나, 융합주 F7, F10 은 각각 4.2 와 8.4 units 로 나타났다. 에탄올 내성 시험을 하여 선별한 결과, 가장 우수한 균주인 융합주 7개 중 F7 과 F10 을 선별하였다. 5% 의 녹말이 포함된 발효배양액에서 $30^{\circ}C$ 5일간 배양시 생성된 에탄올 생성은 융합주 F7 과 F10 은 각각 2.0% 와 1.85 이였다.

• BMB Reports
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• 제39권5호
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• pp.642-647
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• 2006

Botulinum neurotoxin A (BoNT/A) has been used therapeutically to treat muscular hypercontractions and sudomotor hyperactivity and it has been reported that BoNT/A might have analgesic properties in headache. PEP-1 peptide is a known carrier peptide that delivers fulll-ength native proteins in vitro and in vivo. In this study, a BoNT/A gene were fused with PEP-1 peptide in a bacterial expression vector to produce a genetic in-frame PEP-1-BoNT/A fusion protein. The expressed and purified PEP-1-BoNT/A fusion proteins were efficiently transduced into cells in a time- and dose-dependent manner when added exogenously in a culture medium. In addition, immuno-histochemical analysis revealed that PEP-1-BoNT/A fusion protein efficiently penetrated into the epidermis as well as the dermis of the subcutaneous layer, when sprayed on mice skin. These results suggest that PEP-1-BoNT/A fusion protein provide an efficient strategy for therapeutic delivery in various human diseases related to this protein.

• BMB Reports
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• 제41권5호
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• pp.408-413
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• 2008

Pyridoxal-5'-phosphate phosphatase (PLPP) catalyzes the dephosphorylation of pyridoxal-5'-phosphate (PLP). A human brain PLPP gene was fused with a PEP-1 peptide and produced a genetic in-frame PEP-1-PLPP fusion protein. The purified PEP-1-PLPP fusion protein was efficiently transduced into PC12 cells in a time- and dose-dependent manner when added exogenously to culture media. Once inside the cells, the transduced PEP-1-PLPP fusion protein was stable for 36 h. The concentration of PLP was markedly decreased by the addition of exogenous PEP-1-PLPP to media pretreated with the vitamin $B_6$ precursors; pyridoxine, pyridoxal kinase and pyridoxine-5'-phosphate oxidase into cells. The results suggest that the transduction of the PEP-1-PLPP fusion protein can be one mode of PLP level regulation, and to replenish this enzyme in the various neurological disorders related to vitamin $B_6$.

• BMB Reports
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• 제43권1호
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• pp.40-45
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• 2010

AMP-activated protein kinase (AMPK) is a heterotrimeric enzyme that plays a central role in cellular metabolic stress. Modulation of nitric oxide (NO) and cyclooxygenase-2 (COX-2) is considered a promising approach for the treatment of inflammation and neuronal diseases. In this study, the AMPK gene was fused in-frame with PEP-1 peptide in a bacterial expression vector to produce a PEP-1-AMPK fusion protein. Expressed and purified PEP-1-AMPK fusion proteins were transduced efficiently into macrophage Raw 264.7 cells in a time- and dose-dependent manner. Furthermore, transduced PEP-1-AMPK fusion protein markedly inhibited LPS-induced iNOS and COX-2 expression. These results suggest that the PEP-1-AMPK fusion protein can be used for the protein therapy of COX-2 and NO-related disorders such as inflammation and neuronal diseases.

• Molecules and Cells
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• 제42권12호
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• pp.893-905
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• 2019

Mitochondria are highly dynamic organelles that constantly undergo fission and fusion processes that closely related to their function. Disruption of mitochondrial dynamics has been demonstrated in acute kidney injury (AKI), which could eventually result in cell injury and death. Previously, we reported that augmenter of liver regeneration (ALR) alleviates renal tubular epithelial cell injury. Here, we gained further insights into whether the renoprotective roles of ALR are associated with mitochondrial dynamics. Changes in mitochondrial dynamics were examined in experimental models of renal ischemia-reperfusion (IR). In a model of hypoxia-reoxygenation (HR) injury in vitro, dynamin-related protein 1 (Drp1) and mitochondrial fission process protein 1 (MTFP1), two key proteins of mitochondrial fission, were downregulated in the Lv-ALR + HR group. ALR overexpression additionally had an impact on phosphorylation of Drp1 Ser637 during AKI. The inner membrane fusion protein, Optic Atrophy 1 (OPA1), was significantly increased whereas levels of outer membrane fusion proteins Mitofusin-1 and -2 (Mfn1, Mfn2) were not affected in the Lv-ALR + HR group, compared with the control group. Furthermore, the mTOR/4E-BP1 signaling pathway was highly activated in the Lv-ALR + HR group. ALR overexpression led to suppression of HR-induced apoptosis. Our collective findings indicate that ALR gene transfection alleviates mitochondrial injury, possibly through inhibiting fission and promoting fusion of the mitochondrial inner membrane, both of which contribute to reduction of HK-2 cell apoptosis. Additionally, fission processes are potentially mediated by promoting tubular cell survival through activating the mTOR/4E-BP1 signaling pathway.

• BMB Reports
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• 제41권7호
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• pp.537-541
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• 2008

Epilepsy is characterized by the presence of spontaneous episodes of abnormal neuronal discharges and its pathogenic mechanisms remain poorly understood. Recently, we found that the expression of creatine kinase (CK) was markedly decreased in an epilepsy animal model using proteomic analysis. A human CK gene was fused with a HIV-1 Tat peptide to generate an in-frame Tat-CK fusion protein. The purified Tat-CK fusion protein was efficiently transduced into PC12 cells in a time- and dose-dependent manner when added exogenously to culture media. Once inside the cells, the transduced Tat-CK fusion protein was stable for 48 h. Moreover, the Tat-CK fusion protein markedly increased endogenous CK activity levels within the cells. These results suggest that Tat-CK provides a strategy for the therapeutic delivery of proteins in various human diseases including the delivery of CK for potential epilepsy treatment.

• 한국균학회지
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• 제22권1호
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• pp.107-116
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• 1994

원형질체 융합으로 주름버섯목 느타리버섯과 민주름버섯목 잔나비걸상버섯과의 이목간 체세포잡종을 36균주 얻어 꺽쇠연결체 clamp connections있는 2균주와 꺽쇠연결체 없는 3균주의 자실체를 형성하였다. 꺽쇠연결체 없는 융합주는 한천배지나 액체배지에서는 자실체를 얻을 수 없었다. 그러나 톱밥배지에서 균사가 완전히 성장한 후 일정한 광과 온도를 유지한 결과 꺽쇠연결체가 있는 균사가 새로이 성장하였으며 거의 완전하게 다시 성장한 후 원기가 유도되고 자실체가 성숙하였다. 버섯자실체 특성은 느타리버섯과 유사하였으며, 갓 색택은 느타리와는 다소 다르게 나타났다. 3균주의 담자포자로 유전형질의 분리와 유전자재조합을 조사한 결과 꺽쇠연결체 있는 2균주는 비정상적인 분리 현상을 보였는데, 양친에 없는 ane, rib, ane rib 표지를 가진 것이 나타났다. 3종류의 random primer를 이용하여 핵산 연쇄 중합반응 polymerase chain reaction(PCR)에 의한 4개 체세포잡종의 염색체 DNA의 다형화현상을 조사한 결과 느타리친과 유사한 양상을 가졌으나 비양친의 밴드를 나타내었으며 primer #87, #125의 1.2kbp, 0.6kbp에서 각각 뚜렷하게 구분되었다. 4개 체세포잡종의 미토콘드리아 DNA의 제한효소 EcoR1과 HindIII의 절단결과 2균주는 느타리와 동일하였으며 2균주는 양친과 다른 양상을 나타내었다. 원형질체 융합주, 융합주 자실체로부터의 조직배양주, 융합주의 담자포자 발아주, 양친주를 포함하여 총16균주를 등전점 전기영동으로 동위효소 esterase를 분석한 결과 융합주 6균주중 5균주는 느타리와 유사하였으며 1균주는 양친과 전혀 다른 새로운 밴드양상으로 이들 모두 양친과 뚜렷이 구분되었다. 융합주와 자실체 조직배양주는 밴드양상이 거의 유사하였으나 $F^2$는 다소 차이가 나타났다.

• Biotechnology and Bioprocess Engineering:BBE
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• 제4권1호
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• pp.59-62
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• 1999

The recombinant bacteria strain DPD2540, containing a fabA::luxCDABE fusion, was used to detect the toxicity of various chemicals in this study. Membrane damaging agents such as phenol, ethanol, and cerulenin induced a rapid bioluminescent response from this strain. Other toxic agents, such as DNA-damaging or oxidative-damaging chemicals, showed a delayed bioluminescent response in which the maximum peak appeared over 150 min after induction. This strain was also tested for measurement of toxicity in field samples such as wastewater and river water effluents.

• Journal of Microbiology and Biotechnology
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• 제11권1호
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• pp.164-166
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• 2001

To examine the characteristics of the recombinant thin aggregative fimbriae of Salmonella, the AgfA subunit gene was amplified from Salmonella enteritidis using a PCR. The maltose binding protein (MBP)-AgfA fusion protein was overproduced in E. coli and purified. The secondary structure of AgfA was then elucidated from the difference CD spectra. An estimation of the secondary structure of AgfA using the self-consistent method revealed a mostly ${\beta}-sheet$ structure.

• 한국잠사학회:학술대회논문집
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• 한국잠사학회 2005년도 제48회 춘계 학술연구 발표회
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• pp.34-34
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• 2005