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| Content Provider | World Health Organization (WHO)-Global Index Medicus |
|---|---|
| Author | Von Lintig, Johannes Che, Tao Shi, Wuxian Kiser, Philip D. Golczak, Marcin Sui, Xuewu Palczewski, Krzysztof Carey, Paul R. |
| Description | Author Affiliation: Sui X ( From the Departments of Pharmacology and.) |
| Abstract | Carotenoid cleavage enzymes (CCEs) constitute a group of evolutionarily related proteins that metabolize a variety of carotenoid and non-carotenoid substrates. Typically, these enzymes utilize a non-heme iron center to oxidatively cleave a carbon-carbon double bond of a carotenoid substrate. Some members also isomerize specific double bonds in their substrates to yield cis-apocarotenoid products. The apocarotenoid oxygenase from Synechocystis has been hypothesized to represent one such member of this latter category of CCEs. Here, we developed a novel expression and purification protocol that enabled production of soluble, native ACO in quantities sufficient for high resolution structural and spectroscopic investigation of its catalytic mechanism. High performance liquid chromatography and Raman spectroscopy revealed that ACO exclusively formed all-trans products. We also found that linear polyoxyethylene detergents previously used for ACO crystallization strongly inhibited the apocarotenoid oxygenase activity of the enzyme. We crystallized the native enzyme in the absence of apocarotenoid substrate and found electron density in the active site that was similar in appearance to the density previously attributed to a di-cis-apocarotenoid intermediate. Our results clearly demonstrated that ACO is in fact a non-isomerizing member of the CCE family. These results indicate that careful selection of detergent is critical for the success of structural studies aimed at elucidating structures of CCE-carotenoid/retinoid complexes. |
| ISSN | 00219258 |
| e-ISSN | 1083351X |
| Journal | Journal of Biological Chemistry |
| Issue Number | 18 |
| Volume Number | 289 |
| Language | English |
| Publisher | American Society for Biochemistry and Molecular Biology (United States) |
| Publisher Date | 2014-05-02 |
| Publisher Place | United States |
| Access Restriction | Open |
| Subject Keyword | Bacterial Proteins Metabolism Carotenoids Isomerases Oxygenases Chemistry Genetics Biocatalysis Drug Effects Chromatography, High Pressure Liquid Crystallography, X-Ray Kinetics Polyethylene Glycols Pharmacology Retinaldehyde Spectrum Analysis, Raman Synechococcus Enzymology Research Support, N.I.H., Extramural Research Support, U.S. Gov't, Non-P.H.S. Biochemistry Molecular Biology |
| Content Type | Text |
| Resource Type | Article |
| Subject | Cell Biology Biochemistry Molecular Biology |
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