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33 results on '"Kwak, Geun-Hee"'

11. Structural and Biochemical Analysis of Mammalian Methionine Sulfoxide Reductase B2

Author

Aachmann, Finn L., Kwak, Geun-Hee, Conte, Rebecca Del, Kim, Hwa-Young, Gladyshev, Vadim N., and Dikiy, Alexander

Subjects
Models, Molecular, Mice, Binding Sites, Catalytic Domain, Methionine Sulfoxide Reductases, Animals, Hydrogen-Ion Concentration, Nuclear Magnetic Resonance, Biomolecular, Article, Catalysis, Protein Structure, Secondary
Abstract

Methionine sulfoxide reductases are antioxidant enzymes that repair oxidatively damaged methionine residues in proteins. Mammals have three members of the methionine-R-sulfoxide reductase family, including cytosolic MsrB1, mitochondrial MsrB2, and endoplasmic reticulum MsrB3. Here, we report the solution structure of reduced Mus musculus MsrB2 using high resolution nuclear magnetic resonance (NMR) spectroscopy. MsrB2 is a β-strand rich globular protein consisting of eight antiparallel β-strands and three N-terminal α-helical segments. The latter secondary structure elements represent the main structural difference between mammalian MsrB2 and MsrB1. Structural comparison of mammalian and bacterial MsrB structures indicates that the general topology of this MsrB family is maintained and that MsrB2 more resembles bacterial MsrBs than MsrB1. Structural and biochemical analysis supports the catalytic mechanism of MsrB2 that, in contrast to MsrB1, does not involve a resolving cysteine (Cys). pH dependence of catalytically relevant residues in MsrB2 was accessed by NMR spectroscopy and the pK(a) of the catalytic Cys162 was determined to be 8.3. In addition, the pH-dependence of MsrB2 activity showed a maximum at pH 9.0, suggesting that deprotonation of the catalytic Cys is a critical step for the reaction. Further mobility analysis showed a well-structured N-terminal region, which contrasted with the high flexibility of this region in MsrB1. Our study highlights important structural and functional aspects of mammalian MsrB2 and provides a unifying picture for structure-function relationships within the MsrB protein family.

Published
2011

13. Structural and Kinetic Analysis of Free Methionine-R-sulfoxide Reductase from Staphylococcus aureus: CONFORMATIONAL CHANGES DURING CATALYSIS AND IMPLICATIONS FOR THE CATALYTIC AND INHIBITORY MECHANISMS*

Author

Bong, Seoung Min, Kwak, Geun-Hee, Moon, Jin Ho, Lee, Ki Seog, Kim, Hong Seok, Kim, Hwa-Young, and Chi, Young Min

Subjects
Protein Structure and Folding
Abstract

Free methionine-R-sulfoxide reductase (fRMsr) reduces free methionine R-sulfoxide back to methionine, but its catalytic mechanism is poorly understood. Here, we have determined the crystal structures of the reduced, substrate-bound, and oxidized forms of fRMsr from Staphylococcus aureus. Our structural and biochemical analyses suggest the catalytic mechanism of fRMsr in which Cys102 functions as the catalytic residue and Cys68 as the resolving Cys that forms a disulfide bond with Cys102. Cys78, previously thought to be a catalytic Cys, is a non-essential residue for catalytic function. Additionally, our structures provide insights into the enzyme-substrate interaction and the role of active site residues in substrate binding. Structural comparison reveals that conformational changes occur in the active site during catalysis, particularly in the loop of residues 97–106 containing the catalytic Cys102. We have also crystallized a complex between fRMsr and isopropyl alcohol, which acts as a competitive inhibitor for the enzyme. This isopropyl alcohol-bound structure helps us to understand the inhibitory mechanism of fRMsr. Our structural and enzymatic analyses suggest that a branched methyl group in alcohol seems important for competitive inhibition of the fRMsr due to its ability to bind to the active site.

Published
2010

20. Methionine sulfoxide reductase B3 deficiency stimulates heme oxygenase-1 expression via ROS-dependent and Nrf2 activation pathways.

Author

Kwak, Geun-Hee, Kim, Ki Young, and Kim, Hwa-Young

Subjects
*METHIONINE sulfoxide reductase, *HEME oxygenase genetics, *GENE expression, *LEUCINE zippers, *REACTIVE oxygen species, *ACETYLCYSTEINE, *ENDOPLASMIC reticulum
Abstract

Methionine sulfoxide reductase B3 (MsrB3), which is primarily found in the endoplasmic reticulum (ER), is an important protein repair enzyme that stereospecifically reduces methionine- R -sulfoxide residues. We previously found that MsrB3 deficiency arrests the cell cycle at the G 1 /S stage through up-regulation of p21 and p27. In this study, we report a critical role of MsrB3 in gene expression of heme oxygenase-1 (HO-1), which has an anti-proliferative effect associated with p21 up-regulation. Depletion of MsrB3 elevated HO-1 expression in mammalian cells, whereas MsrB3 overexpression had no effect. MsrB3 deficiency increased cellular reactive oxygen species (ROS), particularly in the mitochondria. ER stress, which is associated with up-regulation of HO-1, was also induced by depletion of MsrB3. Treatment with N -acetylcysteine as an ROS scavenger reduced augmented HO-1 levels in MsrB3-depleted cells. MsrB3 deficiency activated Nrf2 transcription factor by enhancing its expression and nuclear import. The activation of Nrf2 induced by MsrB3 depletion was confirmed by increased expression levels of its other target genes, such as γ-glutamylcysteine ligase. Taken together, these data suggest that MsrB3 attenuates HO-1 induction by inhibiting ROS production, ER stress, and Nrf2 activation. [ABSTRACT FROM AUTHOR]

Published
2016
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33. Methionine sulfoxide reductase B3 protects from endoplasmic reticulum stress in Drosophila and in mammalian cells

Author

Kwak, Geun-Hee, Lim, Do-Hwan, Han, Jee Yun, Lee, Young Sik, and Kim, Hwa-Young

Subjects
*METHIONINE sulfoxide, *ENDOPLASMIC reticulum, *PSYCHOLOGICAL stress, *DROSOPHILA, *THAPSIGARGIN, *TUNICAMYCIN, *HOMEOSTASIS, *GENE expression
Abstract

Abstract: Methionine sulfoxide reductase B3A (MsrB3A), which catalyzes the stereospecific reduction of methionine-R-sulfoxide to methionine, is localized to the endoplasmic reticulum (ER). Here, we report a critical role of the ER-targeted MsrB3 in protection against ER stress in Drosophila and in mammalian cells. Flies overexpressing human MsrB3A exhibited significantly increased resistance to ER stress induced by dithiothreitol. These flies also showed slightly enhanced resistance to tunicamycin-induced ER stress. In addition, overexpression of MsrB3A in mammalian cells increased resistance to dithiothreitol- and thapsigargin-induced ER stresses. However, MsrB3A overexpression had no effect on the resistance to tunicamycin-induced ER stress. Knockdown of MsrB3A in mammalian cells led to a significant decrease in the resistance to thapsigargin-induced ER stress, but had no effects on the resistance to either dithiothreitol- or tunicamycin-induced ER stress. Collectively, our data provide evidence that the ER-type of MsrB3 plays an important role in protection against ER stress, suggesting that MsrB3 may be involved in the regulation of ER homeostasis. [Copyright &y& Elsevier]

Published
2012
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