Large-scale interaction profiling of PDZ domains through proteomic peptide-phage display using human and viral phage peptidomes

Ylva Ivarsson; Roland Arnold; Megan McLaughlinSatra NimRakesh JoshiDebashish RayBernard LiuJoan TeyraTony PawsonJason MoffatShawn Shun-Cheng LiSachdev S. SidhuPhilip M. Kim

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Large-scale interaction profiling of PDZ domains through proteomic peptide-phage display using human and viral phage peptidomes

机译：Large-scale interaction profiling of PDZ domains through proteomic peptide-phage display using human and viral phage peptidomes

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摘要

The human proteome contains a plethora of short linear motifs (SLiMs) that serve as binding interfaces for modular protein domains. Such interactions are crucial for signaling and other cellular processes, but are difficult to detect because of their low to moderate affinities. Here we developed a dedicated approach, proteomic peptide-phage display (ProP-PD), to identify domain–SLiM interactions. Specifically, we generated phage libraries containing all human and viral C-terminal peptides using custom oligonucleotide microarrays. With these libraries we screened the nine PSD-95/ Dlg/ZO-1 (PDZ) domains of human Densin-180, Erbin, Scribble, and Disks large homolog 1 for peptide ligands. We identified several known and putative interactions potentially relevant to cellular signaling pathways and confirmed interactions between fulllength Scribble and the target proteins β-PIX, plakophilin-4, and guanylate cyclase soluble subunit α-2 using colocalization and coimmunoprecipitation experiments. The affinities of recombinant Scribble PDZ domains and the synthetic peptides representing the C termini of these proteins were in the 1- to 40-μM range. Furthermore, we identified several well-established host–virus protein– protein interactions, and confirmed that PDZ domains of Scribble interact with the C terminus of Tax-1 of human T-cell leukemia virus with micromolar affinity. Previously unknown putative viral protein ligands for the PDZ domains of Scribble and Erbin were also identified. Thus, we demonstrate that our ProP-PD libraries are useful tools for probing PDZ domain interactions. The method can be extended to interrogate all potential eukaryotic, bacterial, and viral SLiMs and we suggest it will be a highly valuable approach for studying cellular and pathogen–host protein–protein interactions.

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《Proceedings of the National Academy of Sciences of the United States of America》 |2014年第7期|2542-2547|共6页
作者
Ylva Ivarsson; Roland Arnold; Megan McLaughlinSatra NimRakesh JoshiDebashish RayBernard LiuJoan TeyraTony PawsonJason MoffatShawn Shun-Cheng LiSachdev S. SidhuPhilip M. Kim;
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作者单位

Donnelly Centre, University of Toronto, Toronto, ON, Canada M5S 3E1;

Department of Biochemistry, Western University, London, ON, Canada N6A 5C1;

Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, ON, Canada M5G 1X5;

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收录信息美国《科学引文索引》(SCI);美国《生物学医学文摘》(MEDLINE);美国《化学文摘》(CA);
原文格式 PDF
正文语种英语
中图分类自然科学总论;
关键词
Large-scale; proteomic; peptidomes;

Large-scale interaction profiling of PDZ domains through proteomic peptide-phage display using human and viral phage peptidomes

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