Screening For Small Molecule Inhibitors Of p27Kip1 Degradation
| dc.contributor.author | Ooi, Li Ching | |
| dc.date.accessioned | 2019-06-26T06:39:51Z | |
| dc.date.available | 2019-06-26T06:39:51Z | |
| dc.date.issued | 2012-07 | |
| dc.description.abstract | In a majority of human cancers, the cyclin-dependent kinase (CDK) inhibitor p27Kip1 is commonly found to be deregulated due to proteolysis by the ubiquitin-proteasome pathway involving the SCFSkp2 E3 ligase. Although proteasome inhibitors act to stabilize p27Kip1, small molecules agents inhibiting the degradation of p27Kip1 by the function of SCFSkp2 E3 ligase may contribute to the development of a specific targeted drug that could prove to be more efficacious with less side effects. In this work, we have developed a high-throughput screening system based on phosphorylation dependent protein-protein interaction using the baculovirus protein expression system. This system has provided an increased throughput in screening of large libraries of compounds in RIKEN Natural Products Depository (NPDepo) and has facilitated the identification of small molecule inhibitors of p27Kip1 degradation. In the baculovirus protein expression system, S-phase kinase-associated protein 2 (Skp2) tagged with fluorescent monomeric Azami Green (mAG) and CDK subunit 1 (Cks1) (mAGSkp2-Cks1) complex was constructed. The binding of mAGSkp2-Cks1 complex to p27Kip1 phosphopeptides in a phosphorylation dependent manner was confirmed by spectrofluorometry. With this approach, we have screened approximately 20,000 low molecular weight compounds stored in RIKEN NPDepo and have identified two small molecule compounds namely; linichlorin A and gentian violet that were shown to inhibit the interaction between mAGSkp2-Cks1 and p27Kip1 phosphopeptides. Further studies have shown that the compounds inhibited the in vitro ubiquitination of p27Kip1. | en_US |
| dc.identifier.uri | http://hdl.handle.net/123456789/8401 | |
| dc.language.iso | en | en_US |
| dc.publisher | Universiti Sains Malaysia | en_US |
| dc.subject | Screening for small molecule inhibitors | en_US |
| dc.subject | of p27Kip1 degradation | en_US |
| dc.title | Screening For Small Molecule Inhibitors Of p27Kip1 Degradation | en_US |
| dc.type | Thesis | en_US |
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