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Crystal structure of the ferredoxin reductase component of carbazole 1,9a-dioxygenase from Janthinobacterium sp. J3
Yuji Ashikawa1, Zui Fujimoto2, Kengo Inoue3
1Agro-Biotechnology Research Center, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan.
Insights
Crystal structures of carbazole 1,9a-dioxygenase reductase (CARDO-R) reveal key conformational changes affecting flavin adenine dinucleotide (FAD) binding. These insights into electron transfer mechanisms are crucial for understanding Rieske nonheme iron oxygenase function.
Area of Science:
- Biochemistry
- Structural Biology
- Enzymology
Background:
- Carbazole 1,9a-dioxygenase (CARDO) is a Rieske nonheme iron oxygenase (RO) essential for carbazole degradation.
- ROs are classified into five subclasses based on their components and redox centers.
- CARDO comprises an oxygenase, ferredoxin (CARDO-F), and ferredoxin reductase (CARDO-R).
Purpose of the Study:
- To elucidate the structural basis of electron transfer in class III CARDO-R from Janthinobacterium sp. J3 (CARDO-RJ3).
- To investigate the role of flavin adenine dinucleotide (FAD) binding and conformational changes in CARDO-R function.
- To understand the interaction between CARDO-R and its redox partners.
Main Methods:
- X-ray crystallography to resolve two types of CARDO-RJ3 crystal structures (type I and type II).
- Structural superimposition to identify differences between the two crystal forms.
- Molecular docking simulations to predict NADH and CARDO-F interactions with CARDO-RJ3.
Main Results:
- Two distinct crystal structures of CARDO-RJ3 were determined, revealing conformational differences, particularly in the FAD-binding domain and C-terminus.
- Type II structure lacked FAD, showing significant domain movement and altered FAD-binding site.
- Docking simulations suggested that C-terminal residue shifts facilitate electron transfer by positioning NADH close to FAD and indicated favorable complex formation with CARDO-F via electrostatic and shape complementarity.
Conclusions:
- Structural plasticity of CARDO-RJ3, especially concerning FAD binding, is critical for its function in electron transport.
- Distinct domain arrangements compared to other reductases highlight specific adaptations for interaction with their cognate ferredoxins.
- The findings provide a structural basis for understanding electron transfer mechanisms in Rieske nonheme iron oxygenases.
Abstract:
Carbazole 1,9a-dioxygenase (CARDO), which consists of an oxygenase component and the electron-transport components ferredoxin (CARDO-F) and ferredoxin reductase (CARDO-R), is a Rieske nonheme iron oxygenase (RO). ROs are classified into five subclasses (IA, IB, IIA, IIB and III) based on their number of constituents and the nature of their redox centres. In this study, two types of crystal structure (type I and type II) were resolved of the class III CARDO-R from Janthinobacterium sp. J3 (CARDO-RJ3). Superimposition of the type I and type II structures revealed the absence of flavin adenine dinucleotide (FAD) in the type II structure along with significant conformational changes to the FAD-binding domain and the C-terminus, including movements to fill the space in which FAD had been located. Docking simulation of NADH into the FAD-bound form of CARDO-RJ3 suggested that shifts of the residues at the C-terminus caused the nicotinamide moiety to approach the N5 atom of FAD, which might facilitate electron transfer between the redox centres. Differences in domain arrangement were found compared with RO reductases from the ferredoxin-NADP reductase family, suggesting that these differences correspond to differences in the structures of their redox partners ferredoxin and terminal oxygenase. The results of docking simulations with the redox partner class III CARDO-F from Pseudomonas resinovorans CA10 suggested that complex formation suitable for efficient electron transfer is stabilized by electrostatic attraction and complementary shapes of the interacting regions.
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