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Rat liver thioredoxin and thioredoxin reductase: purification and characterization
Abstract
A reproducible scheme has been developed for the preparation of rat liver thioredoxin and thioredoxin reductase (EC 1.6.4.5) by using assays based on reduction of insulin and 5,5'-dithiobis(2-nitrobenzoic acid), respectively. Both proteins were purified to homogeneity, as judged from polyacrylamide gel electrophoresis. Thioredoxin had a molecular weight of 12 000 and contained about 110 amino acids including 4 half-cystines and an NH2-terminal valine. Peptide maps of reduced and carboxymethylated thioredoxin showed that the protein had the active center sequence -Cys-Gly-Pro-Cys-Lys-Met- characteristic of thioredoxins also from procaryotes. Prolonged air oxidation of fully reduced thioredoxin created inactive, aggregated disulfide-containing molecules. Thioredoxin reductase showed a subunit molecular weight of 58 000 and a native molecular weight of 116 000. The enzyme was highly specific for NADPH with a Km of 6 microM. It contained FAD as prosthetic group and was sensitive to inhibition by arsenite. Thioredoxin reductase had a Km of 2.5 microM for rat and calf liver thioredoxin and a Kcat of 3000 min-1.
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Cited by
Schafer, F. Q., & Buettner, G. R. (2001) · Free Radical Biology and Medicine · doi:10.1016/s0891-5849(01)00480-4 · Cited by 4,587
Lee, S.-R., Kwon, K.-S., Kim, S.-R., & Rhee, S. G. (1998) · Journal of Biological Chemistry · doi:10.1074/jbc.273.25.15366 · Cited by 969
Targeting thioredoxin reductase is a basis for cancer therapy by arsenic trioxide
Lu, J., Chew, E.-H., & Holmgren, A. (2007) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.0701549104 · Cited by 526
Self-citationChae, H. Z., Kim, H. J., Kang, S. W., & Rhee, S. G. (1999) · Diabetes Research and Clinical Practice · doi:10.1016/s0168-8227(99)00037-6 · Cited by 390
Redox Signaling Mediated by Thioredoxin and Glutathione Systems in the Central Nervous System
Ren, X., Zou, L., Zhang, X., Branco, V., Wang, J., Carvalho, C., Holmgren, A., & Lu, J. (2017) · Antioxidants & Redox Signaling · doi:10.1089/ars.2016.6925 · Cited by 361
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Metadata
- Authors
- Luthman, M., & Holmgren, A.
- Venue
- Biochemistry
- Publisher
- American Chemical Society (ACS)
- Published
- 1982-12-01
- Volume
- 21
- Issue
- 26
- Pages
- 6628-6633
- ISSN
- 0006-2960 (ISSN-L)
- 1520-4995
- OpenAlex
- W1992533564
- Cited by
- 602
- References
- 0
- FWCI
- 2.29
- Type
- journal article
- Language
- en
- Open access
- closed
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- Luthman, M., & Holmgren, A. (1982). Rat liver thioredoxin and thioredoxin reductase: purification and characterization. Biochemistry. https://doi.org/10.1021/bi00269a003
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