Please use this identifier to cite or link to this item: doi:10.22028/D291-28652
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Title: Characterization of cytochrome P450 CYP109E1 from Bacillus megaterium as a novel vitamin D3 hydroxylase
Author(s): Abdulmughni, Ammar
Jóźwik, Ilona K.
Putkaradze, Natalia
Brill, Elisa
Zapp, Josef
Thunnissen, Andy-Mark W. H.
Hannemann, Frank
Bernhardt, Rita
Language: English
Title: Journal of Biotechnology
Volume: 243
Startpage: 38
Endpage: 47
Publisher/Platform: Elsevier
Year of Publication: 2017
Publikation type: Journal Article
Abstract: In this study the ability of CYP109E1 from Bacillus megaterium to metabolize vitamin D3 (VD3) was investigated. In an in vitro system using bovine adrenodoxin reductase (AdR) and adrenodoxin (Adx4-108), VD3 was converted by CYP109E1 into several products. Furthermore, a whole-cell system in B. megaterium MS941 was established. The new system showed a conversion of 95% after 24h. By NMR analysis it was found that CYP109E1 catalyzes hydroxylation of VD3 at carbons C-24 and C-25, resulting in the formation of 24(S)-hydroxyvitamin D3 (24S(OH)VD3), 25-hydroxyvitamin D3 (25(OH)VD3) and 24S,25-dihydroxyvitamin D3 (24S,25(OH)2VD3). Through time dependent whole-cell conversion of VD3, we identified that the formation of 24S,25(OH)2VD3 by CYP109E1 is derived from VD3 via the intermediate 24S(OH)VD3. Moreover, using docking analysis and site-directed mutagenesis, we identified important active site residues capable of determining substrate specificity and regio-selectivity. HPLC analysis of the whole-cell conversion with the I85A-mutant revealed an increased selectivity towards 25-hydroxylation of VD3 compared with the wild type activity, resulting in an approximately 2-fold increase of 25(OH)VD3 production (45mgl-1day-1) compared to wild type (24.5mgl-1day-1).
DOI of the first publication: 10.1016/j.jbiotec.2016.12.023
URL of the first publication: https://doi.org/10.1016/j.jbiotec.2016.12.023
Link to this record: hdl:20.500.11880/27695
http://dx.doi.org/10.22028/D291-28652
ISSN: 1873-4863
0168-1656
Date of registration: 6-Sep-2019
Faculty: NT - Naturwissenschaftlich- Technische Fakultät
Department: NT - Biowissenschaften
Collections:UniBib – Die Universitätsbibliographie

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