Reconstructing a Flavodoxin Oxidoreductase with Early Amino Acids
Abstract
:1. Introduction
2. Results and Discussion
2.1. Reconstruction of the Flavodoxin Mutant
2.2. Expression and Purification of the Flavodoxin Mutant
2.3. Spectral Assay
2.4. Redox Potentials and Dissociation Constant of the Mutant Flavodoxin
3. Experimental Section
3.1. Expression and Purification of the Mutant Flavodoxin
3.2. Preparation of Apoflavodoxin and Holoflavodoxin
3.3. Spectral Assay
3.4. Determination of Redox Potentials and Dissociation Constant
4. Conclusions
Acknowledgments
Conflict of Interest
References
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Amino acid position | 6 | 7 | 16 | 36 | 54 | 57 | 70 | 71 | 86 | 89 | 91 | 96 | 97 | 100 | 120 | 127 |
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Wild-type | Y | W | M | F | C | M | F | F | F | Y | W | W | M | W | M | C |
Mutant | V | V | L | V | A | V | L | L | V | V | V | V | L | V | V | V |
Secondary structure a | s | s | h | s | h | l | h | h | s | s | s | h | h | h | l | s |
Absorption maxima (nm) | Redox potentials Em,7 (mV) | Kd,ox (μM) | ΔGb,ox (kcal/mol) | Kd,hq (μM) | ΔGb,hq (kcal/mol) | |
---|---|---|---|---|---|---|
FMN | 266, 373, 445 a | −207 a | – | – | – | – |
Wild-type | 272, 378, 446 b | −244 b | 0.00047 b | −12.7 b | 8.1 b | −11.0 b |
Mutant-type | 274, 376, 450 | −360 | 2.92 | −7.52 | 43,604 | −1.85 |
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Lu, M.-F.; Ji, H.-F.; Li, T.-X.; Kang, S.-K.; Zhang, Y.-J.; Zheng, J.-F.; Tian, T.; Jia, X.-S.; Lin, X.-M.; Zhang, H.-Y. Reconstructing a Flavodoxin Oxidoreductase with Early Amino Acids. Int. J. Mol. Sci. 2013, 14, 12843-12852. https://doi.org/10.3390/ijms140612843
Lu M-F, Ji H-F, Li T-X, Kang S-K, Zhang Y-J, Zheng J-F, Tian T, Jia X-S, Lin X-M, Zhang H-Y. Reconstructing a Flavodoxin Oxidoreductase with Early Amino Acids. International Journal of Molecular Sciences. 2013; 14(6):12843-12852. https://doi.org/10.3390/ijms140612843
Chicago/Turabian StyleLu, Ming-Feng, Hong-Fang Ji, Ting-Xuan Li, Shou-Kai Kang, Yue-Jie Zhang, Jue-Fei Zheng, Tian Tian, Xi-Shuai Jia, Xing-Ming Lin, and Hong-Yu Zhang. 2013. "Reconstructing a Flavodoxin Oxidoreductase with Early Amino Acids" International Journal of Molecular Sciences 14, no. 6: 12843-12852. https://doi.org/10.3390/ijms140612843