Plk1-PBD and checked for good hydrogen bond interactions with the five key residues. This resulted in the identification of 526 compounds with good hydrogen bonding. Figure. 6 represents the binding orientation of one hit compound within the Plk1-PBD and also how well the compound fits into Hypo1. To further narrow down the candidate list we placed an extra filter based on the pose and a docking score greater than 60. This resulted in the identification of 93 high confidence compounds likely to inhibit the Plk1-PBD. Interestingly, these compounds have diverse scaffolds that are able to satisfy the geometric constraints on Hypo1 to form similar interactions. Indicating that multiple avenues can be taken to develop therapeutics targeting the Plk1-PBD. The recent interest in developing inhibitors to the Plk1-PBD necessitates a comprehensive analysis of the Plk1-PBD-ligand interaction. Here, we have successfully developed a consensus structure-based pharmacophore model that describes the Plk1PBD-ligand interaction. This structure-based pharmacophore model was integrated with virtual screening and molecular docking approaches to identify 93 potentially novel Plk1 inhibitors, which meet AMDET and Rule of five properties. The testing of these 93 compounds in vitro, with a Plk1-PBD-substrate binding assay, indicated that 
most of the 93 compounds had Plk1-PBD inhibitory activity and that Chemistry_28272 was the most potent compound with an IC50 of 37 mM. Chemistry_28272 represents a new class of Plk1-PBD inhibitors and could serve as a lead compound for further therapeutic development. A major pathway of intracellular protein degradation involves the proteasome, a multi-subunit enzyme complex that resides in the cytosol and nucleus. Proteins destined for degradation, usually by the covalent addition of ubiquitin, are transported into the interior of the proteasome where they encounter the active protease subunits. There are three active subunits: beta 1; beta 2; and beta 5. The proteasome cleaves proteins into peptides typically 3�C25 residues long, and these peptides are usually further degraded into amino acids by a variety of cellular enzymes such as oligoendopeptidases, tripeptidyl peptidase 2, and aminopeptidases. A small percentage of the peptides produced by the proteasome are transported into the endoplasmic PD 0332991 reticulum and incorporated into major histocompatibility complex class I proteins, which present the peptides on the cell surface. Although many proteasome degradation products are rapidly destroyed by aminopeptidases, mass spectrometry based peptidomic studies detected a large number of protein-derived peptides in animal tissues and cell lines. Only a small portion of the peptides detected in the peptidomic studies were derived from the most abundant or most unstable cellular proteins, suggesting that these peptides did not merely reflect protein turnover. Recently, several studies have found that intracellular peptides are functional and influence signal transduction as well as other cellular processes. In an effort to identify the source of the intracellular peptides, previous studies AMN107 treated SH-SY5Y cells and/or HEK293T cells with proteasome inhibitors and examined the effect on the cellular peptidome. One study involved the proteasome inhibitor epoxomicin, an irreversible inhibitor that potently blocks the beta 5 site and also inhibits the beta 2 site at higher concentrations. Most, although not all of the peptides that required cleavage at hydrophobic sites were reduced by treatment with either low or high concentrations of epoxomicin, consistent with the hypothesis that the proteasome was responsible for production of these peptides.