Generation of specific inhibitors of SUMO-1- and SUMO-2/3-mediated protein-protein interactions using Affimer (Adhiron) technology
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Because protein-protein interactions underpin most biological processes, developing tools that target them to understand their function or to inform the development of therapeutics is an important task. SUMOylation is the posttranslational covalent attachment of proteins in the SUMO family (SUMO-1, SUMO-2, or SUMO-3), and it regulates numerous cellular pathways. SUMOylated proteins are recognized by proteins with SUMO-interaction motifs (SIMs) that facilitate noncovalent interactions with SUMO. We describe the use of the Affimer system of peptide display for the rapid isolation of synthetic binding proteins that inhibit SUMO-dependent protein-protein interactions mediated by SIMs both in vitro and in cells. Crucially, these synthetic proteins did not prevent SUMO conjugation either in vitro or in cell-based systems, enabling the specific analysis of SUMO-mediated protein-protein interactions. Furthermore, through structural analysis and molecular modeling, we explored the molecular mechanisms that may underlie their specificity in interfering with either SUMO-1–mediated interactions or interactions mediated by either SUMO-2 or SUMO-3. Not only will these reagents enable investigation of the biological roles of SUMOylation, but the Affimer technology used to generate these synthetic binding proteins could also be exploited to design or validate reagents or therapeutics that target other protein-protein interactions.
Hughes , D J , Tiede , C , Penswick , N , Ah-San Tang , A , Trinh , C H , Mendal , U , Zajac , K Z , Gaule , T , Howell , G , Edwards , T A , Duan , J , Feyfant , E , McPhereson , M J , Tomlinson , D C & Whitehouse , A 2017 , ' Generation of specific inhibitors of SUMO-1- and SUMO-2/3-mediated protein-protein interactions using Affimer (Adhiron) technology ' , Science Signaling , vol. 10 , no. 505 , eaaj2005 . https://doi.org/10.1126/scisignal.aaj2005
Copyright © 2017, the Author(s). This work has been made available online in accordance with the publisher’s policies. This is the author created, accepted version manuscript following peer review and may differ slightly from the final published version. The final published version of this work is available at https://doi.org/10.1126/scisignal.aaj2005
DescriptionThis work was funded in part by a Leeds Cancer Research UK (CRUK) Development Fund and The Wellcome Trust (ISSF) (DJH), The Wellcome Trust (089330), BBSRC (BB/K000306/1 and BB/M006557/1) (AW) and The University of Leeds through Biomedical Health Research Centre support for the Leeds BioScreening Technology Group (MJM, DCT).
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