• KSBB Journal
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• 제24권3호
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• pp.279-286
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• 2009

방선균은 다양한 생리활성 물질을 이차대사산물로 생산하는 산업적으로 매우 유용한 미생물이다. 이에 따라 많은 연구진들이 방선균에 대한 분자생물학적 연구와 산업적 이용에 대한 연구들을 수행하고 있다. 방선균 중에서도 S. avermitilis는 강력한 구충효과가 있는 avermectin을 생산하지만, 또한 포유동물 세포의 미토콘드리아에서 산화적 인산화반응을 억제하는 oligomycin도 함께 생성된다. 따라서 S. avermitilis에서 oligomycin의 생성을 제거시키기 위하여 oligomycin synthetase gene을 disruption 시키기 위한 연구를 수행하였다. 이를 위하여 S. avermitilis로부터 cloning 한 oligomycin synthetase gene (olmA5)의 중앙부분에 apramycin resistance gene을 끼워 넣어 integration vector로 구축한 후에 S. avermitilis의 chromosomal DNA와의 homologous recombination에 의하여 olmA5 gene의 disruption을 유도하였다. Disruption mutants (olmA5::apra)는 PCR을 통해 olmA5 gene의 위치에 apramycin resistance gene이 존재하는 것으로 확인하였고, 또한 HPLC 분석을 통해 oligomycin 생합성이 완전히 제거된 것임을 확인하였다. 그러나 disruption mutant (olmA5::apra)를 이용하여 avermectin 만을 생산할 수 있었으나, avermectin의 생산량에는 거의 변화가 없었다. 이러한 mutants는 산업적으로 avermectin을 생산하기 위한 균주 개량의 훌륭한 source가 될 수 있을 것이다.

• Molecules and Cells
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• 제39권3호
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• pp.211-216
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• 2016

CYP107W1 from Streptomyces avermitilis is a cytochrome P450 enzyme involved in the biosynthesis of macrolide oligomycin A. A previous study reported that CYP107W1 regioselectively hydroxylated C12 of oligomycin C to produce oligomycin A, and the crystal structure of ligand free CYP107W1 was determined. Here, we analyzed the structural properties of the CYP107W1-oligomycin A complex and characterized the functional role of the Trp178 residue in CYP107W1. The crystal structure of the CYP107W1 complex with oligomycin A was determined at a resolution of $2.6{\AA}$. Oligomycin A is bound in the substrate access channel on the upper side of the prosthetic heme mainly by hydrophobic interactions. In particular, the Trp178 residue in the active site intercalates into the large macrolide ring, thereby guiding the substrate into the correct binding orientation for a productive P450 reaction. A Trp178 to Gly mutation resulted in the distortion of binding titration spectra with oligomycin A, whereas binding spectra with azoles were not affected. The Gly178 mutant's catalytic turnover number for the 12-hydroxylation reaction of oligomycin C was highly reduced. These results indicate that Trp178, located in the open pocket of the active site, may be a critical residue for the productive binding conformation of large macrolide substrates.

• 한국미생물·생명공학회지
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• 제23권2호
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• pp.170-177
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• 1995

To search and develop the compounds exhibiting antifungal activities against the mycelial phase of Candida albicans, approximately 2,900 microorganisms isolated from soil were examined for antifungal activity. Among them, a strain with preferential activity against the mycelial phase of Candida albicans was isolated and identified as Streptomyces sp. A393. Isolation and purification of compounds A393 showing antifungal activity against the mycelial phase of C. albicans were performed using XAD-7 column chromatography, silica gel chromatography, preparative thin- layer silica gel chromatography, and HPLC. The molecular weights of compounds isolated from Streptomyces sp. A393 were determined as 774, 790, 804 and 820. These compounds appeared to have a structure of macrolide antibiotics, oligomycin A, B, C and E. Especially, oligomycin E, which is formerly reported to have no antifungal activity, showed antifungal activity against the mycelial phase of Candida albicans.

• 대한지역사회영양학회:학술대회논문집
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• 대한지역사회영양학회 1998년도 추계학술대회 및 제3차 정기총회
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• pp.91-91
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• 1998

• Preventive Nutrition and Food Science
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• 제5권4호
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• pp.230-233
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• 2000

The present work was designed to determine whether change in fluidity of the mitochondrial membrane affects mitochondrial {TEX}$F_{1}${/TEX}{TEX}$F_{0}${/TEX}ATPase characteristics in NIDDM-prone BHE/Cdb rat. Isolated mitochondria fom BHE/Cdb rat fed a 6% coconut oil or corn oil were functionally tested by an analysis of its respiration and the coupling of this process to ATP synthesis in presence of oligomycin, a specific inhibitor of oxidative phosphorylation (OXPHOS), that binds to the {TEX}$F_{1}${/TEX}{TEX}$F_{0}${/TEX}ATPase. Mitochondria from rats fed coconut oil were more responsive to the inhibitory action of oligomycin with respect to state 3 respiration, respiratory control (RC) ratio and ADP:P (P/O) ratio than were mitochondria from rats fed corn oil. In state 3 respiration, mitochondria from rats fed coconut oil consumed less oxygen than did mitochondria from rats fed corn oil. RC ratio was lower in the mitochondria from rats fed coconut oil than was mitochondria from rats fed corn oil. In P/O ratio, the mitochondria from rats fed coconut oil had a lower P/O ratio than did mitochondria from rats fed corn oil. The data showed that the chang influidity of the mitochondrial membrane by dietary fat affected mitochondrial {TEX}$F_{1}${/TEX}{TEX}$F_{0}${/TEX}ATPase characteristics. The present study on diet differences in {TEX}$F_{1}${/TEX}{TEX}$F_{0}${/TEX}ATPase characteristics provides considerable insight into the role diets play in the control of mitochondrial function, expecially OXPHOS in NIDDM with mitochondrial defects.

• IMMUNE NETWORK
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• 제22권2호
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• pp.18.1-18.15
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• 2022

Dysfunction of mitochondrial metabolism is implicated in cellular injury and cell death. While mitochondrial dysfunction is associated with lung injury by lung inflammation, the mechanism by which the impairment of mitochondrial ATP synthesis regulates necroptosis during acute lung injury (ALI) by lung inflammation is unclear. Here, we showed that the impairment of mitochondrial ATP synthesis induces receptor interacting serine/threonine kinase 3 (RIPK3)-dependent necroptosis during lung injury by lung inflammation. We found that the impairment of mitochondrial ATP synthesis by oligomycin, an inhibitor of ATP synthase, resulted in increased lung injury and RIPK3 levels in lung tissues during lung inflammation by LPS in mice. The elevated RIPK3 and RIPK3 phosphorylation levels by oligomycin resulted in high mixed lineage kinase domain-like (MLKL) phosphorylation, the terminal molecule in necroptotic cell death pathway, in lung epithelial cells during lung inflammation. Moreover, the levels of protein in bronchoalveolar lavage fluid (BALF) were increased by the activation of necroptosis via oligomycin during lung inflammation. Furthermore, the levels of ATP5A, a catalytic subunit of the mitochondrial ATP synthase complex for ATP synthesis, were reduced in lung epithelial cells of lung tissues from patients with acute respiratory distress syndrome (ARDS), the most severe form of ALI. The levels of RIPK3, RIPK3 phosphorylation and MLKL phosphorylation were elevated in lung epithelial cells in patients with ARDS. Our results suggest that the impairment of mitochondrial ATP synthesis induces RIPK3-dependent necroptosis in lung epithelial cells during lung injury by lung inflammation.

• The Korean Journal of Physiology
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• 제8권2호
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• pp.59-66
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• 1974

In recent years much interesting information about the mechanism of the $Na^+-K^+$ activated ATPase has been obtained from investigation of the $K^+-activated$ phosphatase activity which appears to be catalysed by the same enzyme. Also several studies have indicated that a $K^+-activated p-nitrophenylphosphatase activity is intimately related to the ATPase activity. And then the exact relation of p-nitrophenylphosphatase activity to $Na^+-K^+$ ATPase activity is not known. The effects of some ions and drugs on the p-nitrophenylphosphatase activity of the rat brain were investigated and the results were summarized as follows. 1. The p-nitrophenylphosphatase was stimulated markedly by low concentrations of $K^+$, while the activity was activated slightly in the presence of $Na^+$ and oligomycin. 2. Addition of both ATP and $Na^+$ caused a remarkable increase in the activity of the $K^+-dependent$ phosphatase at low concentrations of $K^+$. 3. In the presence of $Na^+$ and low concentrations of $K^+$, oligomycin activated the p-nitrophenylphosphatase. 4. O1igomycin inhibited the stimulation of the enzyme activity caused by $Na^{+}+ATP$. 5. Ouabain inhibited the $K^+-dependent$ p-nitrophenylphosphatase activity more in the presence of ATP and $Na^+$ than in their absence. 6. Quinidine inhibited both $Na^+-K^+$ ATPase and p-nitrophenylphosphatase. These inhibitory effects of the drug were partially antagonized by increasing $K^+$ concentrations. The sensitivity of the $K^+-dependent$ p-nitrophenylphosphatase to quinidine was greater than the that of $Na^+-K^+$ ATPase.

• 한국균학회지
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• 제15권4호
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• pp.224-230
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• 1987

표고버섯의 미토콘드리아는 설탕밀도 선형기울기 원심분리법으로 정제하여, 광감응성 mitochondrial ATPase의 유기물 효과, 광증감제 효과 및 파장 변화에 따른 $K^+$ 이온의 유입 효과를 실험하였다. 1. 이 효소는 10m mol dithiothreitol 및 0.1m mol quinacrine에 의하여 각각 139% 및 128%의 활성도를 증가시켰다. 2. $100\;{\mu}g$의 oligomycin과 1m mol phlorizin은 이 효소의 활성을 각각 48% 및 45% 억제시켰다. 3. 광증감제인 0.1m mol phenazine methosulfate는 이 효소의 활성도를 36% 촉진시켰다. 4. $K^+$ 이온 유입 효과의 최적 파장은 690nm였고, 이때의 최적 pH 및 최적 온도는 각각 7.2 및 $55^{\circ}C$였다.

• 한국균학회지
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• 제17권2호
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• pp.99-104
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• 1989

1. 최적광조사 조건, 470 nm에서 15초 동안 빛을 조사한 미토콘드리아성 ATP synthase는 1mmole DCPI에 의하여 그 활성이 85% 증가되었다. 2. 1mmole DNP, $10\;{\mu}mole$ HHQNO 및 $100\;{\mu}g/ml$의 oligomycin은 이 효소의 활성을 각각 61, 41% 및 12% 억제시켰다. 3. 최적 조건의 빛에 의한 $Fe^{3+}$ 및 $Fe^{2+}$ 이온의 유입효과에서, 5mmole $Fe^{3+}$ 및 0.5mmole $Fe^{2+}$ 이온에 의하여 이 효소 활성이 각각 14% 및 12% 증가되었다. 4. 광증감제인 phenazine methosulfate의 존재하에서 470nm의 빛을 조사한 후의 효소의 활성도가 크게 증가되는 것으로 보아, 미토콘드리아성 ATP synthase내에 미지의 흡광물질이 존재함을 추정할 수 있었다.

• Molecules and Cells
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• 제23권3호
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• pp.363-369
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• 2007

Mitochondria play a central role in energy-generating processes and may be involved in the regulation of channels and receptors. Here we investigated TRPM7, an ion channel and functional kinase, and its regulation by mitochondria. Proton ionophores such as CCCP elicited a rapid decrease in outward TRPM7 whole-cell currents but a slight increase in inward currents with pipette solutions containing no MgATP. With pipette solutions containing 3 mM MgATP, however, CCCP increased both outward and inward TRPM7 currents. This effect was reproducible and fully reversible, and repeated application of CCCP yielded similar decreases in current amplitude. Oligomycin, an inhibitor of $F_1/F_O$-ATP synthase, inhibited outward whole-cell currents but did not affect inward currents. The respiratory chain complex I inhibitor, rotenone, and complex III inhibitor, antimycin A, were without effect as were kaempferol, an activator of the mitochondrial $Ca^{2+}$ uniporter, and ruthenium red, an inhibitor of the mitochondrial $Ca^{2+}$ uniporter. These results suggest that the inner membrane potential (as regulated by proton ionophores) and the $F_1/F_O$-ATP synthase of mitochondria are important in regulating TRPM7 channels.