Cyclic Cascade in Metabolic Regulation

P . Boon Chock 한국미생물생명공학회 ( 구 한국산업미생물학회 ) 1993 한국미생물생명공학회 학술발표초록집 Vol.1993 No.1

Protein Glutathionylation in the Regulation of Peroxiredoxins: A Family of Thiol-Specific Peroxidases That Function As Antioxidants, Molecular Chaperones, and Signal Modulators

Chae, Ho Zoon,Oubrahim, Hammou,Park, Ji Won,Rhee, Sue Goo,Chock, P. Boon Mary Ann Liebert 2012 Antioxidants & redox signaling Vol.16 No.6

Glutathionylation of Peroxiredoxin I Induces Decamer to Dimers Dissociation with Concomitant Loss of Chaperone Activity

Park, Ji Won,Piszczek, Grzegorz,Rhee, Sue Goo,Chock, P. Boon American Chemical Society 2011 Biochemistry Vol.50 No.15

<P>Reversible protein glutathionylation, a redox-sensitive regulatory mechanism, plays a key role in cellular regulation and cell signaling. Peroxiredoxins (Prxs), a family of peroxidases that is involved in removing H<SUB>2</SUB>O<SUB>2</SUB> and organic hydroperoxides, are known to undergo a functional change from peroxidase to molecular chaperone upon overoxidation of its catalytic cysteine. The functional change is caused by a structural change from low molecular weight oligomers to high molecular weight complexes that possess molecular chaperone activity. We reported earlier that Prx I can be glutathionylated at three of its cysteine residues, Cys52, -83, and -173 [<name name-style='western'>Park</name><etal>et al.</etal><x xml:space='preserve'> (</x>2009<x xml:space='preserve'>) </x>J. Biol. Chem., 284<x xml:space='preserve'>, </x>23364]. In this study, using analytical ultracentrifugation analysis, we reveal that glutathionylation of Prx I, WT, or its C52S/C173S double mutant shifted its oligomeric status from decamers to a population consisting mainly of dimers. Cys83 is localized at the putative dimer−dimer interface, implying that the redox status of Cys83 may play an important role in stabilizing the oligomeric state of Prx I. Studies with the Prx I (C83S) mutant show that while Cys83 is not essential for the formation of high molecular weight complexes, it affects the dimer−decamer equilibrium. Glutathionylation of the C83S mutant leads to accumulation of dimers and monomers. In addition, glutathionylation of Prx I, both the WT and C52S/C173S mutants, greatly reduces their molecular chaperone activity in protecting citrate synthase from thermally induced aggregation. Together, these results reveal that glutathionylation of Prx I promotes changes in its quaternary structure from decamers to smaller oligomers and concomitantly inactivates its molecular chaperone function.</P><P><B>Graphic Abstract</B> <IMG SRC='http://pubs.acs.org/appl/literatum/publisher/achs/journals/content/bichaw/2011/bichaw.2011.50.issue-15/bi101373h/production/images/medium/bi-2010-01373h_0006.gif'></P>

Deglutathionylation of 2-Cys peroxiredoxin is specifically catalyzed by sulfiredoxin.

Park, Ji Won,Mieyal, John J,Rhee, Sue Goo,Chock, P Boon American Society for Biochemistry and Molecular Bi 2009 The Journal of biological chemistry Vol.284 No.35

<P>Reversible protein glutathionylation plays a key role in cellular regulation and cell signaling and protects protein thiols from hyperoxidation. Sulfiredoxin (Srx), an enzyme that catalyzes the reduction of Cys-sulfinic acid derivatives of 2-Cys peroxiredoxins (2-Cys Prxs), has been shown to catalyze the deglutathionylation of actin. We show that deglutathionylation of 2-Cys Prx, a family of peroxidases, is specifically catalyzed by Srx. Using the ubiquitously expressed member of 2-Cys Prx, Prx I, we revealed the following. (i) Among its four Cys residues, Cys(52), Cys(83), and Cys(173) can be glutathionylated in vitro. Deglutathionylation with Cys mutants showed that Cys(83) and Cys(173) were preferentially catalyzed by Srx, with glutathionylated Srx as the reaction intermediate, whereas glutaredoxin I was more favorable for deglutathionylating Cys(52). (ii) Studies using site-directed mutagenesis coupled with binding and deglutathionylation activities revealed that Pro(174) and Pro(179) of Prx I and Tyr(92) of Srx are essential for both activities. Furthermore, relative to glutaredoxin I, Srx exhibited negligible deglutathionylation activity for glutathionylated cysteine and glutathionylated BSA. These results indicate that Srx is specific for deglutathionylating Prx I due to its favorable affinity for Prx I. To assess the biological relevance of these observations, we showed that Prx I is glutathionylated in A549 and HeLa cells under modest levels of H(2)O(2). In addition, the level of glutathionylated Prx I was substantially elevated in small interfering RNA-mediated Srx-knocked down cells, whereas the reverse was observed in Srx-overexpressing cells. However, glutathionylation of Prx V, not known to bind to Srx, was not affected by the change in Srx expression levels.</P>

Novel protective mechanism against irreversible hyperoxidation of peroxiredoxin: Nalpha-terminal acetylation of human peroxiredoxin II.

Seo, Jae Ho,Lim, Jung Chae,Lee, Duck-Yeon,Kim, Kyung Seok,Piszczek, Grzegorz,Nam, Hyung Wook,Kim, Yu Sam,Ahn, Taeho,Yun, Chul-Ho,Kim, Kanghwa,Chock, P Boon,Chae, Ho Zoon American Society for Biochemistry and Molecular Bi 2009 The Journal of biological chemistry Vol.284 No.20

<P>Peroxiredoxins (Prxs) are a group of peroxidases containing a cysteine thiol at their catalytic site. During peroxidase catalysis, the catalytic cysteine, referred to as the peroxidatic cysteine (C(P)), cycles between thiol (C(P)-SH) and disulfide (-S-S-) states via a sulfenic (C(P)-SOH) intermediate. Hyperoxidation of the C(P) thiol to its sulfinic (C(P)-SO(2)H) derivative has been shown to be reversible, but its sulfonic (C(P)-SO(3)H) derivative is irreversible. Our comparative study of hyperoxidation and regeneration of Prx I and Prx II in HeLa cells revealed that Prx II is more susceptible than Prx I to hyperoxidation and that the majority of the hyperoxidized Prx II formation is reversible. However, the hyperoxidized Prx I showed much less reversibility because of the formation of its irreversible sulfonic derivative, as verified with C(P)-SO(3)H-specific antiserum. In an attempt to identify the multiple hyperoxidized spots of the Prx I on two-dimensional PAGE analysis, an N-acetylated Prx I was identified as part of the total Prx I using anti-acetylated Lys antibody. Using peptidyl-Asp metalloendopeptidase (EC 3.4.24.33) peptide fingerprints, we found that N(alpha)-terminal acetylation (N(alpha)-Ac) occurred exclusively on Prx II after demethionylation. N(alpha)-Ac of Prx II blocks Prx II from irreversible hyperoxidation without altering its affinity for hydrogen peroxide. A comparative study of non-N(alpha)-acetylated and N(alpha)-terminal acetylated Prx II revealed that N(alpha)-Ac of Prx II induces a significant shift in the circular dichroism spectrum and elevation of T(m) from 59.6 to 70.9 degrees C. These findings suggest that the structural maintenance of Prx II by N(alpha)-Ac may be responsible for preventing its hyperoxidation to form C(P)-SO(3)H.</P>

Structural and biochemical analyses reveal ubiquitin C-terminal hydrolase-L1 as a specific client of the peroxiredoxin II chaperone

Lee, Sang Pil,Park, Chan Mi,Kim, Kyung Seok,Kim, Eunji,Jeong, Moonkyung,Shin, Ji-Young,Yun, Chul-Ho,Kim, Kanghwa,Chock, P. Boon,Chae, Ho Zoon Elsevier 2018 Archives of biochemistry and biophysics Vol.640 No.-

<P><B>Abstract</B></P> <P>Peroxiredoxins (Prxs) play dual roles as both thiol-peroxidases and molecular chaperones. Peroxidase activity enables various intracellular functions, however, the physiological roles of Prxs as chaperones are not well established. To study the chaperoning function of Prx, we previously sought to identify heat-induced Prx-binding proteins as the clients of a Prx chaperone. By using His-tagged Prx I as a bait, we separated ubiquitin C-terminal hydrolase-L1 (UCH-L1) as a heat-induced Prx I binding protein from rat brain crude extracts. Protein complex immunoprecipitation with HeLa cell lysates revealed that both Prx I and Prx II interact with UCH-L1. However, Prx II interacted considerably more favorably with UCH-L1 than Prx I. Prx II exhibited more effective molecular chaperone activity than Prx I when UCH-L1 was the client. Prx II interacted with UCH-L1 through its C-terminal region to protect UCH-L1 from thermal or oxidative inactivation. We found that chaperoning via interaction through C-terminal region (specific-client chaperoning) is more efficient than that involving oligomeric structural change (general-client chaperoning). Prx II binds either thermally or oxidatively unfolding early intermediates of specific clients and thereby shifted the equilibrium towards their native state. We conclude that this chaperoning mechanism provides a very effective and selective chaperoning activity.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Prx II interacts with UCH-L1 through its C-terminal region. </LI> <LI> Prx II effectively protects UCH-L1 against thermal or oxidative inactivation. </LI> <LI> Oligomeric structural changes are not prerequisite for this chaperoning mechanism. </LI> <LI> This novel chaperoning mechanism is named as specific-client chaperoning. </LI> </UL> </P>