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학술연구/단체지원/교육 등 연구자 활동을 지속하도록 DBpia가 지원하고 있어요.
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논문 기본 정보
- 자료유형
- 학술저널
- 저자정보
- 저널정보
- 한국미생물생명공학회 Journal of Microbiology and Biotechnology Journal of Microbiology and Biotechnology 제27권 제5호
- 발행연도
- 2017.1
- 수록면
- 983 - 989 (7page)
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초록· 키워드
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NADPH-P450 reductase (NPR) transfers electrons from NADPH to cytochrome P450 and heme oxygenase enzymes to support their catalytic activities. This protein is localized within the endoplasmic reticulum membrane and utilizes FMN, FAD, and NADPH as cofactors.
Although NPR is essential toward enabling the biochemical and pharmacological analyses of P450 enzymes, its production as a recombinant purified protein requires a series of tedious efforts and a high cost due to the use of NADP+ in the affinity chromatography process. In the present study, the rat NPR clone containing a 6× Histidine-tag (NPR-His) was constructed and heterologously expressed. The NPR-His protein was purified using Ni2+-affinity chromatography, and its functional features were characterized. A single band at 78 kDa was observed from SDS-PAGE and the purified protein displayed a maximum absorbance at 455 nm, indicating the presence of an oxidized flavin cofactor. Cytochrome c and nitroblue tetrazolium were reduced by purified NPR-His in an NADPH-dependent manner. The purified NPR-His successfully supported the catalytic activities of human P450 1A2 and 2A6 and fungal CYP52A21, yielding results similar to those obtained using conventional purified rat reductase. This study will facilitate the use of recombinant NPR-His protein in the various fields of P450 research.
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참고문헌 (20)
참고문헌 신청
Guengerich FP / 1998 / Purification of cytochromes P450. Rat and human hepatic forms / Methods Mol. Biol 107 : 35 ~ 53
Guengerich FP / 2001 / Principles and Methods of Toxicology / Taylor &Francis : 1625 ~ 1687
Guengerich FP / 2009 / Measurement of cytochrome P450 and NADPH-cytochrome P450 reductase / Nat. Protoc 4 : 1245 ~ 1251
Han S / 2015 / Functional characterization of CYP107W1 from Streptomyces avermitilis and biosynthesis of macrolide oligomycin A / Arch. Biochem. Biophys 575 : 1 ~ 7
Hanna IH / 1998 / Role of the alanine at position 363 of cytochrome P450 2B2in influencing the NADPH- and hydroperoxide-supported activities / Arch. Biochem. Biophys 350 : 32
Guengerich FP / 2001 / Principles and Methods of Toxicology / Taylor &Francis : 1625 ~ 1687
Guengerich FP / 2009 / Measurement of cytochrome P450 and NADPH-cytochrome P450 reductase / Nat. Protoc 4 : 1245 ~ 1251
Han S / 2015 / Functional characterization of CYP107W1 from Streptomyces avermitilis and biosynthesis of macrolide oligomycin A / Arch. Biochem. Biophys 575 : 1 ~ 7
Hanna IH / 1998 / Role of the alanine at position 363 of cytochrome P450 2B2in influencing the NADPH- and hydroperoxide-supported activities / Arch. Biochem. Biophys 350 : 32
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