Biophysical and structural analyses of the interaction between the SHANK1 PDZ domain and an internal SLiM.

IF 4.4 3区 生物学 Q2 BIOCHEMISTRY & MOLECULAR BIOLOGY Biochemical Journal Pub Date : 2024-07-17 DOI:10.1042/BCJ20240126
Yue Li, Chi H Trinh, Amanda Acevedo-Jake, Diana Gimenez, Stuart L Warriner, Andrew J Wilson
{"title":"Biophysical and structural analyses of the interaction between the SHANK1 PDZ domain and an internal SLiM.","authors":"Yue Li, Chi H Trinh, Amanda Acevedo-Jake, Diana Gimenez, Stuart L Warriner, Andrew J Wilson","doi":"10.1042/BCJ20240126","DOIUrl":null,"url":null,"abstract":"<p><p>The PDZ (Postsynaptic density protein-95[PSD-95]/Discs-large) domain, prevalent as a recognition module, has attracted significant attention given its ability to specifically recognize ligands with consensus motifs (also termed PDZ binding motifs [PBMs]). PBMs typically bear a C-terminal carboxylate as a recognition handle and have been extensively characterized, whilst internal ligands are less well known. Here we characterize a short linear motif (SLiM) - EESTSFQGP - as an internal PBM based on its strong binding affinity towards the SHANK1 PDZ domain (SHANK1656-762 hereafter referred to as SHANK1). Using the acetylated analogue Ac-EESTSFQGP-CONH2 as a competitor for the interaction of SHANK1 with FAM-Ahx-EESTSFQGP-CONH2 or a typical fluorophore-labelled C-terminal PBM - GKAP - FITC-Ahx-EAQTRL-COOH - the internal SLiM was demonstrated to show comparable low-micromolar IC50 by competition fluorescent anisotropy. To gain further insight into the internal ligand interaction at the molecular level, we obtained the X-ray co-crystal structure of the Ac-EESTSFQGP-CONH2/SHANK1 complex and compared this to the Ac-EAQTRL-COOH/SHANK1 complex. The crystallographic studies reveal that the SHANK1 backbones for the two interactions overlap significantly. The main structural differences were shown to result from the flexible loops which reorganize to accommodate the two PBMs with distinct lengths and terminal groups. In addition, the two C-terminal residues Gly and Pro in Ac-EESTSFQGP-CONH2 were shown not to participate in interaction with the target protein, implying further truncation and structural modification using peptidomimetic approaches on this sequence may be feasible. Taken together, the SLiM Ac-EESTSFQGP-CONH2 holds potential as an internal ligand for targeting SHANK1.</p>","PeriodicalId":8825,"journal":{"name":"Biochemical Journal","volume":" ","pages":"945-955"},"PeriodicalIF":4.4000,"publicationDate":"2024-07-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11346428/pdf/","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Biochemical Journal","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1042/BCJ20240126","RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"BIOCHEMISTRY & MOLECULAR BIOLOGY","Score":null,"Total":0}
引用次数: 0

Abstract

The PDZ (Postsynaptic density protein-95[PSD-95]/Discs-large) domain, prevalent as a recognition module, has attracted significant attention given its ability to specifically recognize ligands with consensus motifs (also termed PDZ binding motifs [PBMs]). PBMs typically bear a C-terminal carboxylate as a recognition handle and have been extensively characterized, whilst internal ligands are less well known. Here we characterize a short linear motif (SLiM) - EESTSFQGP - as an internal PBM based on its strong binding affinity towards the SHANK1 PDZ domain (SHANK1656-762 hereafter referred to as SHANK1). Using the acetylated analogue Ac-EESTSFQGP-CONH2 as a competitor for the interaction of SHANK1 with FAM-Ahx-EESTSFQGP-CONH2 or a typical fluorophore-labelled C-terminal PBM - GKAP - FITC-Ahx-EAQTRL-COOH - the internal SLiM was demonstrated to show comparable low-micromolar IC50 by competition fluorescent anisotropy. To gain further insight into the internal ligand interaction at the molecular level, we obtained the X-ray co-crystal structure of the Ac-EESTSFQGP-CONH2/SHANK1 complex and compared this to the Ac-EAQTRL-COOH/SHANK1 complex. The crystallographic studies reveal that the SHANK1 backbones for the two interactions overlap significantly. The main structural differences were shown to result from the flexible loops which reorganize to accommodate the two PBMs with distinct lengths and terminal groups. In addition, the two C-terminal residues Gly and Pro in Ac-EESTSFQGP-CONH2 were shown not to participate in interaction with the target protein, implying further truncation and structural modification using peptidomimetic approaches on this sequence may be feasible. Taken together, the SLiM Ac-EESTSFQGP-CONH2 holds potential as an internal ligand for targeting SHANK1.

查看原文
分享 分享
微信好友 朋友圈 QQ好友 复制链接
本刊更多论文
SHANK1 PDZ 域与内部 SLiM 之间相互作用的生物物理和结构分析。
PDZ(突触后密度蛋白-95[PSD-95]/Discs-large)结构域是一种识别模块,由于它能够特异性地识别具有共识基调(也称为 PDZ 结合基调 [PBM])的配体,因此备受关注。PBM 通常以 C 端羧酸盐作为识别柄,并已被广泛表征,而内部配体则鲜为人知。在这里,我们根据短线性基团(SLiM)--EESTSFQGP--与 SHANK1 PDZ 结构域(SHANK1656-762,以下简称 SHANK1)的强结合亲和力,将其表征为内部 PBM。使用乙酰化类似物 Ac-EESTSFQGP-CONH2 作为 SHANK1 与 FAM-Ahx-EESTSFQGP-CONH2 或典型的荧光团标记的 C 端 PBM(GKAP-FITC-Ahx-EAQTRL-COOH)相互作用的竞争物,通过竞争荧光各向异性(FA)证明内部 SLiM 显示出相似的低微摩尔 IC50。为了进一步了解内部配体在分子水平上的相互作用,我们获得了 Ac-EESTSFQGP-CONH2/SHANK1 复合物的 X 射线共晶体结构,并将其与 Ac-EAQTRL-COOH/SHANK1 复合物进行了比较。晶体学研究显示,这两种相互作用的 SHANK1 主干有明显重叠。主要的结构差异来自于柔性环,这些柔性环重组以容纳两个具有不同长度和末端基团的 PBM。此外,Ac-EESTSFQGP-CONH2 中的两个 C 端残基 Gly 和 Pro 未参与与目标蛋白的相互作用,这意味着使用拟肽方法对该序列进行进一步截短和结构修饰是可行的。综上所述,SLiM Ac-EESTSFQGP-CONH2 有潜力成为靶向 SHANK1 的内部配体。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
求助全文
约1分钟内获得全文 去求助
来源期刊
Biochemical Journal
Biochemical Journal 生物-生化与分子生物学
CiteScore
8.00
自引率
0.00%
发文量
255
审稿时长
1 months
期刊介绍: Exploring the molecular mechanisms that underpin key biological processes, the Biochemical Journal is a leading bioscience journal publishing high-impact scientific research papers and reviews on the latest advances and new mechanistic concepts in the fields of biochemistry, cellular biosciences and molecular biology. The Journal and its Editorial Board are committed to publishing work that provides a significant advance to current understanding or mechanistic insights; studies that go beyond observational work using in vitro and/or in vivo approaches are welcomed. Painless publishing: All papers undergo a rigorous peer review process; however, the Editorial Board is committed to ensuring that, if revisions are recommended, extra experiments not necessary to the paper will not be asked for. Areas covered in the journal include: Cell biology Chemical biology Energy processes Gene expression and regulation Mechanisms of disease Metabolism Molecular structure and function Plant biology Signalling
期刊最新文献
Divergent roles of DRY and NPxxY motifs in selective activation of downstream signalling by the apelin receptor. Sequence variation in the active site of mobile colistin resistance proteins is evolutionarily accommodated through inter-domain interactions. The uncharacterized protein ZNF200 interacts with PRMT3 and aids its stability and nuclear translocation. Mitigating neuroinflammation in cognitive areas: exploring the impact of HMG-CoA reductase inhibitor. Pyroptotic executioner pore-forming protein gasdermin D forms oligomeric assembly and exhibits amyloid-like attributes that could contribute for its pore-forming function.
×
引用
GB/T 7714-2015
复制
MLA
复制
APA
复制
导出至
BibTeX EndNote RefMan NoteFirst NoteExpress
×
×
提示
您的信息不完整,为了账户安全,请先补充。
现在去补充
×
提示
您因"违规操作"
具体请查看互助需知
我知道了
×
提示
现在去查看 取消
×
提示
确定
0
微信
客服QQ
Book学术公众号 扫码关注我们
反馈
×
意见反馈
请填写您的意见或建议
请填写您的手机或邮箱
已复制链接
已复制链接
快去分享给好友吧!
我知道了
×
扫码分享
扫码分享
Book学术官方微信
Book学术文献互助
Book学术文献互助群
群 号:481959085
Book学术
文献互助 智能选刊 最新文献 互助须知 联系我们:info@booksci.cn
Book学术提供免费学术资源搜索服务,方便国内外学者检索中英文文献。致力于提供最便捷和优质的服务体验。
Copyright © 2023 Book学术 All rights reserved.
ghs 京公网安备 11010802042870号 京ICP备2023020795号-1