Structural Characterization of KEAP1 R415K to Guide the Design of KEAP1‐Nrf2 Peptide Inhibitors

KEAP1型 氧化应激 化学 转录因子 对接(动物) 结合位点 生物化学 氧化磷酸化 基因 医学 护理部
作者
Gwendolyn Pyeatt,Samantha N. Muellers,Adrian Whitty,Karen N. Allen
出处
期刊:The FASEB Journal [Wiley]
卷期号:35 (S1)
标识
DOI:10.1096/fasebj.2021.35.s1.04626
摘要

Oxidative stress can cause extensive damage to DNA, proteins, and lipids. Thus, long-term oxidative stress has been linked to many diseases including cancer and Alzheimer's disease. The highly-charged binding interface between regulatory protein Kelch-like ECH-associated protein 1 (KEAP1) and transcription factor nuclear erythroid factor 2-like 2 (Nrf2) is a common drug target for diseases related to oxidative stress. In the absence of oxidative stress, KEAP1 binds with high affinity to the DxETGE motif of Nrf2. Oxidative stress causes KEAP1 to release Nrf2, allowing it to translocate to the nucleus and upregulate the transcription of antioxidant enzymes. Our current peptide inhibitors of this protein-protein interaction (PPI) are modeled after the DxETGE motif and have exhibited low cell permeability, prompting the bioisosteric replacement of charged residues. We previously identified Arg415 as the KEAP1 residue with the most significant contribution to binding energy at the KEAP1-Nrf2 interface. It has been observed that the R415A mutation abolishes Nrf2 binding, and so a more conservative KEAP1 R415K variant was characterized to better understand the interactions between this residue and residue Glu79 on Nrf2. The variant was characterized using X-ray crystallography, differential scanning fluorimetry, and the binding was assessed using a competitive fluorescence anisotropy binding assay. This mutation caused a 40-fold decrease in binding affinity, which suggested that retaining the charge alone was not sufficient to restore wild-type binding affinity. KEAP1 R415K was successfully crystallized in 8% w/v polyethylene glycol 20,000, 0.2 M imidazole, and 0.01 M nickel (II) chloride hexahydrate, and the crystals diffracted to a resolution of 2.43 Å. Structures of the unliganded R415K Kelch domain were determined using molecular replacement. Optimization of the crystals toward the structure determination of the R415K variant bound to peptide inhibitors is underway. These liganded structures can then be used to assess the binding interactions between KEAP1 Arg415 and Nrf2 Glu79 and guide future peptide inhibitor optimizations to improve their potency and bioavailability.

科研通智能强力驱动
Strongly Powered by AbleSci AI
更新
PDF的下载单位、IP信息已删除 (2025-6-4)

科研通是完全免费的文献互助平台,具备全网最快的应助速度,最高的求助完成率。 对每一个文献求助,科研通都将尽心尽力,给求助人一个满意的交代。
实时播报
刚刚
朱建军应助科研通管家采纳,获得10
刚刚
李爱国应助科研通管家采纳,获得10
1秒前
香蕉觅云应助科研通管家采纳,获得10
1秒前
1秒前
RC_Wang应助科研通管家采纳,获得10
1秒前
1秒前
共享精神应助Volcano采纳,获得10
5秒前
5秒前
5秒前
ali完成签到,获得积分20
6秒前
han发布了新的文献求助10
7秒前
大个应助无限雨南采纳,获得10
8秒前
丘比特应助幽默尔蓝采纳,获得10
8秒前
EED发布了新的文献求助10
9秒前
9秒前
yy完成签到,获得积分10
10秒前
10秒前
10秒前
李爱国应助十三儿采纳,获得10
11秒前
ali发布了新的文献求助10
12秒前
12秒前
HOME发布了新的文献求助10
14秒前
熊大哥发布了新的文献求助10
16秒前
17秒前
科研马完成签到,获得积分10
18秒前
猫和老鼠发布了新的文献求助10
19秒前
19秒前
20秒前
发篇Sci不过分吧完成签到,获得积分20
20秒前
奥特超曼应助易槐采纳,获得10
22秒前
22秒前
上官若男应助Supreme采纳,获得10
22秒前
熊大哥完成签到,获得积分10
22秒前
22秒前
23秒前
小蘑菇应助威武的手链采纳,获得10
23秒前
可爱千兰完成签到,获得积分10
23秒前
十三儿完成签到,获得积分20
23秒前
24秒前
高分求助中
A new approach to the extrapolation of accelerated life test data 1000
ACSM’s Guidelines for Exercise Testing and Prescription, 12th edition 500
‘Unruly’ Children: Historical Fieldnotes and Learning Morality in a Taiwan Village (New Departures in Anthropology) 400
Indomethacinのヒトにおける経皮吸収 400
Phylogenetic study of the order Polydesmida (Myriapoda: Diplopoda) 370
基于可调谐半导体激光吸收光谱技术泄漏气体检测系统的研究 350
Robot-supported joining of reinforcement textiles with one-sided sewing heads 320
热门求助领域 (近24小时)
化学 材料科学 医学 生物 工程类 有机化学 生物化学 物理 内科学 纳米技术 计算机科学 化学工程 复合材料 遗传学 基因 物理化学 催化作用 冶金 细胞生物学 免疫学
热门帖子
关注 科研通微信公众号,转发送积分 3988920
求助须知:如何正确求助?哪些是违规求助? 3531290
关于积分的说明 11253247
捐赠科研通 3269903
什么是DOI,文献DOI怎么找? 1804830
邀请新用户注册赠送积分活动 882027
科研通“疑难数据库(出版商)”最低求助积分说明 809052