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ELECTROPHORETIC, SPECTRAL, CATALYTIC AND IMMUNOCHEMICAL PROPERTIES OF HIGHLY PURIFIED CYTOCHROME-P-450 FROM SHEEP LUNG
Date
1990-01-01
Author
Adalı, Orhan
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1. 1. Cytochrome P-450LgM2 was purified from sheep lung microsomes in the presence of detergents, Emulgen 913 and cholate. 2. 2. The purification procedure involved the chromatography of the detergent solubilized microsomes on DEAE-cellulose and hydroxylapatite. 3. 3. Cytochrome P-450LgM2 was further purified on second DEAE-cellulose and hydroxylapatite columns. 4. 4. The specific content of the highly purified P-450LgM2 was 16–18 nmol P-450/mg protein and purified 164-fold. 5. 5. The yield was 16% of the initial content in microsomes. 6. 6. The SDS-polyacrylamide slab gel electrophoresis (PAGE) of the purified lung cytochrome P-450LgM2 showed one protein band having the monomer molecular weight of 49,500. 7. 7. The absolute CO-difference spectrum of dithionate-reduced P-450LgM2 gave a peak at 451 nm. 8. 8. When sheep lung cytochrome P-450LgM2 and P-450LM2 purified from liver of phenobarbital (PB)-induced rabbit were subjected to Western Blotting and visualized immunochemically with antiP-450LM2, they showed identical mobilities. 9. 9. P-450LgM2 was found to be very active in N-demethylation of benzphetamine in a reconstituted system containing purified sheep lung reductase and synthetic lipid. 10. 10. Turnover numbers (min−1) for benzphetamine, aniline, ethylmorphine and p-nitrophenol were determined to be 273, 1.2, 15.5 and 1.05, respectively, in a reconstituted microsomal lung monooxygenase system. 11. 11. Spectral, electrophoretic, biocatalytic and immunochemical properties of sheep lung P-450LgM2 were found to be similar to those of P-450 isozyme 2, purified from PB-treated rabbit liver and of rabbit lung microsomes.
Subject Keywords
Biochemistry
URI
https://hdl.handle.net/11511/39472
Journal
INTERNATIONAL JOURNAL OF BIOCHEMISTRY
DOI
https://doi.org/10.1016/0020-711x(90)90234-t
Collections
Department of Biology, Article
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O. Adalı, “ELECTROPHORETIC, SPECTRAL, CATALYTIC AND IMMUNOCHEMICAL PROPERTIES OF HIGHLY PURIFIED CYTOCHROME-P-450 FROM SHEEP LUNG,”
INTERNATIONAL JOURNAL OF BIOCHEMISTRY
, pp. 1433–1432, 1990, Accessed: 00, 2020. [Online]. Available: https://hdl.handle.net/11511/39472.