Crystal Structure of the Kelch Domain of Human Keap1

Li, Xuchu; Zhang, Donna; Hannink, Mark; Beamer, Lesa J.

doi:10.1074/jbc.m410073200

Cited by 202 publications

(191 citation statements)

References 54 publications

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“…These results suggest that the inhibitory regulation effect of OsPPKL1 results from its Kelch domains. The Kelch domain has been reported to represent one β-sheet blade, and several repeats are able to interact to form a β-propeller (17). In rice, LARGER PANICLE (LP) encoding a Kelch repeat-containing F-box protein was reported to play a negative role in regulating plant architecture, particularly panicle architecture (18).…”

Section: Resultsmentioning

confidence: 99%

Rare allele of OsPPKL1 associated with grain length causes extra-large grain and a significant yield increase in rice

Zhang

Wang

Huang

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

423

315

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Grain size and shape are important components determining rice grain yield, and they are controlled by quantitative trait loci (QTLs). Here, we report the cloning and functional characterization of a major grain length QTL, qGL3, which encodes a putative protein phosphatase with Kelch-like repeat domain (OsPPKL1). We found a rare allele qgl3 that leads to a long grain phenotype by an aspartate-to-glutamate transition in a conserved AVLDT motif of the second Kelch domain in OsPPKL1. The rice genome has other two OsPPKL1 homologs, OsPPKL2 and OsPPKL3. Transgenic studies showed that OsPPKL1 and OsPPKL3 function as negative regulators of grain length, whereas OsPPKL2 as a positive regulator. The Kelch domains are essential for the OsPPKL1 biological function. Field trials showed that the application of the qgl3 allele could significantly increase grain yield in both inbred and hybrid rice varieties, due to its favorable effect on grain length, filling, and weight.QTL mapping | positional cloning | Oryza sativa L.

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Section: Resultsmentioning

confidence: 99%

Rare allele of OsPPKL1 associated with grain length causes extra-large grain and a significant yield increase in rice

Zhang

Wang

Huang

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

423

315

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“…Recent reports by several groups have demonstrated that Keap1 functions as a substrate adaptor protein for a Cul3-Rbx1 E3 1 ubiquitin ligase complex (19 -22). The N-terminal BTB domain and central linker region of Keap1 bind Cul3, whereas the C-terminal Kelch domain of Keap1 binds Nrf2 via residues located within loops that extend out from the bottom of the Kelch domain (20,21,23). Under conditions of homeostatic cell growth, Keap1 brings Nrf2 into the Cul3-Rbx1 complex and enables ubiquitin conjugation onto specific lysine residues located within the N-terminal Neh2 domain of Nrf2 (21).…”

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confidence: 99%

Ubiquitination of Keap1, a BTB-Kelch Substrate Adaptor Protein for Cul3, Targets Keap1 for Degradation by a Proteasome-independent Pathway

Zhang

Sun

et al. 2005

Journal of Biological Chemistry

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260

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Keap1 is a BTB-Kelch protein that functions as a substrate adaptor protein for a Cul3-dependent E3 ubiquitin ligase complex. Keap1 targets its substrate, the Nrf2 transcription factor, for ubiquitination and subsequent degradation by the 26 S proteasome. Inhibition of Keap1-dependent ubiquitination of Nrf2 increases steady-state levels of Nrf2 and enables activation of cytoprotective Nrf2-dependent genes. In this report, we demonstrate that Keap1 and three other BTB-Kelch proteins, including GAN1, ENC1, and Sarcosin, are ubiquitinated by a Cul3-dependent complex. Ubiquitination of Keap1 is markedly increased in cells exposed to quinone-induced oxidative stress, occurs in parallel with inhibition of Keap1-dependent ubiquitination of Nrf2, and results in decreased steady-state levels of Keap1, particularly in cells that are unable to synthesize glutathione. Degradation of Keap1 is independent of the 26 S proteasome, because inhibitors of the 26 S proteasome do not prevent loss of Keap1 following exposure of cells to quinone-induced oxidative stress. Our results suggest that a switch from substrate to substrate adaptor ubiquitination is a critical regulatory step that controls steady-state levels of both BTB-Kelch substrate adaptor proteins and their cognate substrates.

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“…Human Keap1 is a 70-kDa cysteine-rich protein (624 amino acids and 27 cysteines), comprising five domains: (i) N-terminal region (NTR), (ii) BTB (broad complex, tramtrack, or bric-a-brac), an evolutionarily conserved protein-protein interaction motif that often dimerizes with other BTB domains (20), (iii) intervening region (IVR), a cysteine-rich region, (iv) double glycine region (DGR), a domain that comprises six Kelch motifs and binds to the Neh2 domain of Nrf2 (21), and (v) C-terminal region (CTR). It is difficult to express soluble Keap1, which is highly prone to oxidation and oligomerizes easily.…”

mentioning

confidence: 99%

Electrophilic tuning of the chemoprotective natural product sulforaphane

Ahn

Hwang

Liu

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

150

125

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Sulforaphane [1-isothiocyanato-4-(methylsulfinyl)butane], a naturally occurring isothiocyanate derived from cruciferous vegetables, is a highly potent inducer of phase 2 cytoprotective enzymes and can protect against electrophiles including carcinogens, oxidative stress, and inflammation. The mechanism of action of sulforaphane is believed to involve modifications of critical cysteine residues of Keap1, which lead to stabilization of Nrf2 to activate the antioxidant response element of phase 2 enzymes. However, the dithiocarbamate functional group formed by a reversible reaction between isothiocyanate of sulforaphane and sulfhydryl nucleophiles of Keap1 is kinetically labile, and such modification in intact cells has not yet been demonstrated. Here we designed sulforaphane analogs with replacement of the reactive isothiocyanate by the more gentle electrophilic sulfoxythiocarbamate group that also selectively targets cysteine residues in proteins but forms stable thiocarbamate adducts. Twenty-four sulfoxythiocarbamate analogs were synthesized that retain the structural features important for high potency in sulforaphane analogs: the sulfoxide or keto group and its appropriate distance to electrophilic functional group. Evaluation in various cell lines including hepatoma cells, retinal pigment epithelial cells, and keratinocytes as well as in mouse skin shows that these analogs maintain high potency and efficacy for phase 2 enzyme induction as well as the inhibitory effect on lipopolysaccharide-induced nitric oxide formation like sulforaphane. We further show in living cells that a sulfoxythiocarbamate analog can label Keap1 on several key cysteine residues as well as other cellular proteins offering new insights into the mechanism of chemoprotection.

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Crystal Structure of the Kelch Domain of Human Keap1

Cited by 202 publications

References 54 publications

Rare allele of OsPPKL1 associated with grain length causes extra-large grain and a significant yield increase in rice

Rare allele of OsPPKL1 associated with grain length causes extra-large grain and a significant yield increase in rice

Ubiquitination of Keap1, a BTB-Kelch Substrate Adaptor Protein for Cul3, Targets Keap1 for Degradation by a Proteasome-independent Pathway

Electrophilic tuning of the chemoprotective natural product sulforaphane

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