| Posttranslational modifications play an important role in regulating cellular process by controlling the biological activity of protein. This dissertation focuses on using one-bead-one compound (OBOC) combinatorial library to study the biological functions of posttranslational modifications., which include (1) discovering substrates of protein tyrosine sulfation, (2) developing peptidomimetic substrates of protein tyrosine kinase, and (3) developing ligand targeting histone modification. Protein tyrosine sulfation, which affects protein-protein interactions, is an ubiquitous post-translational modification catalyzed by isozyme tyrosylprotein sulfotransferases (TPST) 1 and 2 in eukaryotic cells. By screening an OBOC combinatorial peptide library (XXXYXXX, wherein Y = tyrosine, X = all 19 amino acids except cysteine), we have identified a number of peptide substrates for both TPST1 and TPST2. The identified TPST2 peptide substrates show a consensus aspartic acid residue (D) at the -1 position, and they also contain many negatively charged residues in their sequences. A selective peptide to TPST2, WGDYEDD was discovered. BLASTing the sequences of these peptides with protein database enable us to predict and subsequently confirm that complement factor b is a sulfated protein. Second, we designed, synthesized, and screened several peptidomimetic OBOC combinatorial libraries against protein tyrosine kinase, c-Src, which is known to be an oncoprotein and plays an important role in many signal transductions. Followed by a series of structure-activity relationship studies, selective peptidomimetic substrates for c-Src were identified. LC/MS study revealed that a small peptidomimetic compound was able to enter cells and act as a c-Src kinase substrate. This peptidomimetic can serve as a lead compound to develop molecular tracer for PET imaging. Third, we developed several acryloylated peptides targeting H3K4 or H3K9, which can be used to study histone lysine modification, or serve as a labeling tool to mark un-modified lysine, in order to study the physiological functions of H3K4 and H3K9. This is a proof of concept study, which demonstrated that we can design, synthesize and screened an acryloylated OBOC peptide library to develop covalent protein capture reagents that selectively target specific lysine at the histone tail. Furthermore, in collaboration with Dr. Nian-huan Yao, several flavone analogues targeting eEF1A with anti-proliferation activity against breast cancer cells have been developed and their mechanism of action determined. |
