• KSBB Journal
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• 제14권3호
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• pp.259-263
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• 1999

물고기 기름에 대한 Aspergillus oryzae 유래의 Lipolase-100T의 가수분해 특성을 규명하였다. Lipolase-100T는 트리글리세라이드의 1과 3의 위치에 작용하여 이 위치에 결합되어 있는 아실 체인을 유리지방산으로 가수분해시키는 1,3-위채특이성을 보였다. 또한 Lipolase-100T는 물고기 기름을 구성하는 다중불포화지방산 보다 포화지방산을 쉽게 가수분해시키는 특성도 나타내었다. 이 특성으로 인하여 가수분해 시간에 따라서 생성되는 글리세라이드 혼합물내의 다중 불포화 지방산중 n-3 PUFAs인 C16:4, C20:5 및 C22:6의 농도는 모두 증가하였으며 특히 C22:6의 농도증가가 가장 뚜렷하였다. 이 결과로부터 물고기 기름을 구성하는 n-3 PUFAs는 트리글리세라이드의 2번 위치에 결합하고 있음을 알 수 있었다. 지방산의 결합위치특이성과 Lipolase-100T 1,3-위치특이성으로 인하여 Lipolase-100T를 반응물의 0.4 wt%사용하여 물고기 기름을 120시간 가수분해 시켰을 때 생성된 글리세라이드 혼합물내의 n-3 PUFAs농도는 50 wt%까지 상승하였다.

• Applied Biological Chemistry
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• 제23권1호
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• pp.52-57
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• 1980

Trichosporon cutaneum lipase의 작용기작(作用機作) 및 위치특이성(位置特異性)을 thin-layer chromatography 및 densitometer를 사용(使用)하여 검토(檢討)하였던 바 다음과 같은 결과(結果)를 얻었다. 1. 본(本) lipase 지질분해속도(脂質分解速度)는 triolein>diolein>monooleie의 순(順)이었다. 2. 본(本) lipase의 diolein분해속도(分解速度)는 triolein의 그것과 비슷한 정도(程度)로 빨랐다. 3. 본(本) lipase의 monoolein분해속도(分解速度)는 triolein 및 diolein의 그것에 비(比)하여 현저(顯著)히 늦었다. 4. 본(本) lipase는 극(極)히 미약(微弱)한 ${\alpha}$-위치특이성(位置特異性)을 나타내었다.

• Journal of Microbiology and Biotechnology
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• 제9권5호
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• pp.624-630
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• 1999

In order to investigate the position of various fatty acids attached to glycerol and the specificity of Lipolase-100T, hydrolysis of fish oil was carried out with Lipolase-100T derived from Aspergillus oryzae. The amounts of free fatty acids produced from triglyceride, 1,2(2,3)-diglyceride, 1,3-diglyceride, and 2-monoglyceride and conversion rates of 1,2(2,3)-diglyceride to 1,3-diglyceride and 2-monoglyceride to 1(3)-monoglyceride were also calculated. The ratio of 1,2-diglyceride content to 1,3-diglyceride was higher than 70 in the early period of hydrolysis. The fatty acid content of the glyceride mixture after 72 h of hydrolysis was compared with that of fish oil, and it was found that polyunsaturated fatty acids such as C16:4, C20:4 n-3, C20:5 n-3, C21:5 n-3, C22:5 n-3 and C22:6 n-3 were located in the 2-position of glycerol. Material balance of each component in the hydrolysis system was written to obtain a set of simultaneous linear equations. The theoretical quantity of free fatty acids produced from triglyceride, 1,2-diglyceride, 1,3-diglyceride, and monoglyceride, respectively, were calculated by solving the linear equation system. The conversion rate of 1,2(2,3)-diglyceride to 1,3-diglyceride and that of 2-monoglyceride to 1(3)-monoglyceride were also obtained. The results showed that the migration rate of 1,2(2,3)-diglyceride to 1,3-diglyceride was higher than the hydrolysis rate of 1,2(2,3)-diglyceride to 2-monoglyceride and the conversion rate of 2-monoglyceride to 1(3)-monoglyceride was extremely low.

• BMB Reports
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• 제46권3호
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• pp.151-156
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• 2013

Phylogenetic and amino acid sequence analysis indicated that rice allene oxide synthase-1 (OsAOS1) is CYP74, and is clearly distinct from CYP74B, C and D subfamilies. Regio- and stereo-chemical analysis revealed the dual substrate specificity of OsAOS1 for (cis,trans)-configurational isomers of 13(S)- and 9(S)-hydroperoxyoctadecadienoic acid. GC-MS analysis showed that OsAOS1 converts 13(S)- and 9(S)-hydroperoxyoctadecadi(tri)enoic acid into their corresponding allene oxide. UV-Visible spectral analysis of native OsAOS1 revealed a Soret maximum at 393 nm, which shifted to 424 nm with several clean isobestic points upon binding of OsAOS1 to imidazole. The spectral shift induced by imidazole correlated with inhibition of OsAOS1 activity, implying that imidazole may coordinate to ferric heme iron, triggering a heme-iron transition from high spin state to low spin state. The implications and significance of a putative type II ligand-induced spin state transition in OsAOS1 are discussed.

• BMB Reports
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• 제29권4호
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• pp.386-392
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• 1996

A Saccharomyces cerevisiae Protein Tyrosine Phosphatase, PTP1, was expressed from an Escherichia coli expression system and milligram quantities of active PTP1 were purified chromatographically. The substrate specificity of the recombinant PTP1 was probed using synthetic phosphotyrosine-containing peptides corresponding to the regulatory phosphorylation sites of the yeast MAP kinase homologues $Fus3_{176-186}$, $Kss1_{179-189}$, and $Hog1_{170-180}$. Peptide sequences derived from the MAP kinase homologues were chosen arbitrarily as starting points for sequence variation studies even though they are not likely to be candidates for physiological substrates of PTP1. Phosphotyrosyl-$Hog1_{170-180}$ peptide showed a $K_M$ value of 877 ${\mu}M$ and phosphorylated $Kss1_{179-189}$ and $Fus3_{176-186}$ peptides showed lower $K_M$ values of 74 ${\mu}M$ and 51 ${\mu}M$ each. To study the effect of sequence variations of the peptide, amino acids of the undecapeptide $Hog1_{170-180}$ (DPQMTGpYVSTR) were sequentially substituted by an alanine residue. More extensive variations of each amino acid revealed positional importance of each amino acid residue. Based on these results, we derived a peptide sequence (DADEpYDA) that is recognized by PTP1 with an affinity ($K_M$ is 4 ${\mu}M$) significantly higher than that of the peptides derived from the phosphorylation sites of Fus3, Kss1, and Hog1.

• Journal of Microbiology and Biotechnology
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• 제10권4호
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• pp.502-506
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• 2000

In order to determine the position and the content of fatty acids sttached to glycerides and the migration degree of fatty acids in the migration reaction, fish oil was hydroyzed with lipolase-100T which was derived from Aspergillus oryzae. The content of fatty acids in the glyceride mixture was analyzed and compared with that of fish oil. The amounts of fatty acid in a 2-position and the migration degree of the fatty acid in 2,3-DG (diglyceride) and 2-MG (monolyceride) were carefully calculated. The results showed that approximately 95% (w/w) of DHA (docosahexanoic acid) and 65% of EPA(eicosapentaenoic acid) were attached to the 2-position of glycerides in fish oil. Approximately 87% (w/w) of DHA and 75% of EPA remained in 2,3-DG, and 88% of DHA and 65% of EPA in 2-MG were not involved in the migration reaction.

• 한국식품영양과학회지
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• 제38권4호
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• pp.479-485
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• 2009

PMF와 stearic acid를 기질로 하고, TAG의 sn-1,3 위치에 특이적으로 작용하는 lipozyme TLIM을 이용해 항온교반수조($70^{\circ}C$, 180rpm)에서 개질된 PMF(팜중부유)를 얻었다. 효소량을 20 wt%를 사용하였을 경우, 가장 높은 stearic acid 결합율을 보인 PMF와 stearic acid의 몰 비율은 1:2였고, 1:1.5와 비교적 비슷한 함량을 나타내었다. 이에 반해 1:0.5는 가장 낮은 결합율을 보였다. PMF와 stearic acid의 몰 비율 1:2일 때, 효소량을 20 wt% 사용하여 반응한 경우 stearic acid의 함량은 31.6%까지 증가한 반면, 효소량을 10wt% 사용하였을 때는 13.4%까지 증가하여 효소량을 20wt% 사용했을 때보다 2.4배 낮은 함량을 보였다. RP-HPLC 분석결과 반응 기질로 사용된 PMF는 대부분이 POP로 93.27 area%를 차지하였고, 효소 첨가량 20 wt%, PMF와 stearic acid의 몰 비율 1:2로 36 hr 반응한 시료의 TAG 조성은 7 hr 반응했을 때에 비하여 POP가 크게 감소(7 hr: 33.4 area%, 36 hr: 6.21 area%)하였다. 하지만 반응이 진행됨에 따라 크게 증가할 것으로 예측했던 POS(PSO)의 경우 7 hr 반응했을 때보다 2.61% 감소하였다. 반면 7 hr 반응했을 때 보다 PPS는 약 1.9배, PSS는 약 6.7배 증가하는 경향을 보였다. 이로써 lipozyme TLIM으로 반응할 경우 반응시간이 지날수록 acyl migration 현상이 증가하는 것으로 예측되었다. 효소 첨가량 20 wt%, PMF와 stearic acid의 몰 비율 1:2로 36 hr 반응 후 시료의 TAG 중 sn-2 position에는 oleic acid 함량이 5.4%로 낮았고 palmitic acid와 stearic acid의 함량은 각각 50.1%, 43.7%로 나타났다. 이로써 개질된 PMF에서 효소의 sn-1,3 위치 특이성이 나타나지 않은 것으로 예측되었다.