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    Structure, Function, and Inhibition of S100B

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    Author
    Charpentier, Thomas H.
    Advisor
    Weber, David J., Ph.D.
    Date
    2009
    Type
    dissertation
    
    Metadata
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    Abstract
    Aberrant levels of the small dimeric protein S100B have been found in malignant melanoma, renal cell cancer, and astrocytomas. S100B may aid in cancer progression via its interaction with and down regulation of the tumor suppressor p53, in a Ca²⁺ and possibly Zn²⁺ dependent manner. S100B bound to Ca²⁺ undergoes a conformational change exposing a hydrophobic cleft for the p53-S100B interaction. S100B binds to the C-terminus and tetramerization domains (319-393) of p53. Experiments reducing S100B expression via siRNA restores p53 levels in primary malignant melanoma cells. Thus, several small molecules have been identified that bind S100B and inhibit the Ca²⁺-S100B-p53 interaction. One of these small molecules is pentamidine, an FDA approved drug. We have characterized the interaction between Ca²⁺-S100B and pentamidine via nuclear magnetic resonance (NMR) and X-ray crystallography. We obtained crystal structures of pentamidine bound to Zn²⁺-Ca²⁺-S100B. The previously solved NMR structure of Zn²⁺-Ca²⁺-S100B was compared to the X-ray crystal structure solved here. We characterized the Zn²⁺ ligands in each structure to determine if Zn²⁺ binding changed the pentamidine interaction with S100B. A goal of the Weber lab has been to identify small molecules to inhibit the p53-S100B interaction and we have been moderately successful. We have identified three additional small molecules found through screens performed by our lab and the high throughput screening core (UMAB). SBi132, SBi279, and SBi523 (S100B inhibitor ###) were shown to interact with S100B through NMR and X-ray crystallography. Other small molecules derived from pentamidine or SBi132 interact with the "hinge" region of S100B, while other screening molecules were found to covalently bind to cysteine 84 on helix 4 of S100B. To characterize the S100B-p53 protein-protein interaction further, TRTK-12 a peptide derived from the CapZ protein, was used to study the effects of peptide bound to S100B. Surprisingly, the Ca²⁺ coordination for both EF-hands of S100B were not affected by TRTK-12. The X-ray structure of TRTK-12 peptide bound to S100B did show differences in temperature factor. These differences in peptide binding can aid us in identifying inhibitors of the S100B-p53 complex and restore p53 levels in malignant melanoma.
    Description
    University of Maryland, Baltimore. Biochemistry. Ph.D. 2009
    Keyword
    p53
    protein interactions
    S100B
    small molecule
    Calcium
    Genes, p53
    Identifier to cite or link to this item
    http://hdl.handle.net/10713/882
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    Theses and Dissertations School of Medicine
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