A cell permeable bimane-constrained PCNA-interacting peptide.

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dc.contributor.author Horsfall, Aimee J
dc.contributor.author Vandborg, Beth A
dc.contributor.author Kikhtyak, Zoya
dc.contributor.author Scanlon, Denis B
dc.contributor.author Tilley, Wayne D
dc.contributor.author Hickey, Theresa E
dc.contributor.author Bruning, John B
dc.contributor.author Abell, Andrew D
dc.coverage.spatial England
dc.date.accessioned 2022-03-07T01:20:46Z
dc.date.available 2022-03-07T01:20:46Z
dc.date.issued 2021-10
dc.identifier.citation RSC chemical biology 2(5):1499-1508 Oct 2021
dc.identifier.issn 2633-0679
dc.identifier.uri https://hdl.handle.net/2292/58486
dc.description.abstract The human sliding clamp protein known as proliferating cell nuclear antigen (PCNA) orchestrates DNA-replication and -repair and as such is an ideal therapeutic target for proliferative diseases, including cancer. Peptides derived from the human p21 protein bind PCNA with high affinity <i>via</i> a 3<sub>10</sub>-helical binding conformation and are known to shut down DNA-replication. Here, we present studies on short analogues of p21 peptides (143-151) conformationally constrained with a covalent linker between <i>i</i>, <i>i</i> + 4 separated cysteine residues at positions 145 and 149 to access peptidomimetics that target PCNA. The resulting macrocycles bind PCNA with <i>K</i> <sub>D</sub> values ranging from 570 nM to 3.86 μM, with the bimane-constrained peptide <b>7</b> proving the most potent. Subsequent X-ray crystallography and computational modelling studies of the macrocyclic peptides bound to PCNA indicated only the high-affinity peptide <b>7</b> adopted the classical 3<sub>10</sub>-helical binding conformation. This suggests the 3<sub>10</sub>-helical conformation is critical to high affinity PCNA binding, however NMR secondary shift analysis of peptide <b>7</b> revealed this secondary structure was not well-defined in solution. Peptide <b>7</b> is cell permeable and localised to the cell cytosol of breast cancer cells (MDA-MB-468), revealed by confocal microscopy showing blue fluorescence of the bimane linker. The inherent fluorescence of the bimane moiety present in peptide <b>7</b> allowed it to be directly imaged in the cell uptake assay, without attachment of an auxiliary fluorescent tag. This highlights a significant benefit of using a bimane constraint to access conformationally constrained macrocyclic peptides. This study identifies a small peptidomimetic that binds PCNA with higher affinity than previous reported p21 macrocycles, and is cell permeable, providing a significant advance toward development of a PCNA inhibitor for therapeutic applications.
dc.format.medium Electronic-eCollection
dc.language eng
dc.publisher Royal Society of Chemistry (RSC)
dc.relation.ispartofseries RSC chemical biology
dc.rights Items in ResearchSpace are protected by copyright, with all rights reserved, unless otherwise indicated. Previously published items are made available in accordance with the copyright policy of the publisher.
dc.rights.uri https://researchspace.auckland.ac.nz/docs/uoa-docs/rights.htm
dc.rights.uri https://creativecommons.org/licenses/by-nc/3.0/
dc.subject Science & Technology
dc.subject Life Sciences & Biomedicine
dc.subject Physical Sciences
dc.subject Biochemistry & Molecular Biology
dc.subject Chemistry, Multidisciplinary
dc.subject Chemistry
dc.subject INHIBITOR
dc.subject AFFINITY
dc.subject BINDING
dc.subject DOMAIN
dc.subject P21
dc.title A cell permeable bimane-constrained PCNA-interacting peptide.
dc.type Journal Article
dc.identifier.doi 10.1039/d1cb00113b
pubs.issue 5
pubs.begin-page 1499
pubs.volume 2
dc.date.updated 2022-02-15T22:16:39Z
dc.rights.holder Copyright: The author en
pubs.author-url https://www.ncbi.nlm.nih.gov/pubmed/34704055
pubs.end-page 1508
pubs.publication-status Published
dc.rights.accessrights http://purl.org/eprint/accessRights/OpenAccess en
pubs.subtype research-article
pubs.subtype Journal Article
pubs.elements-id 868010
dc.identifier.eissn 2633-0679
dc.identifier.pii d1cb00113b
pubs.online-publication-date 2021

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