title: Screening of protein-protein interaction modulators via sulfo-click kinetic target-guided synthesis.
abstract: Kinetic target-guided synthesis (TGS) and in situ click chemistry are among unconventional discovery strategies having the potential to streamline the development of protein-protein interaction modulators (PPIMs). In kinetic TGS and in situ click chemistry, the target is directly involved in the assembly of its own potent, bidentate ligand from a pool of reactive fragments. Herein, we report the use and validation of kinetic TGS based on the sulfo-click reaction between thio acids and sulfonyl azides as a screening and synthesis platform for the identification of high-quality PPIMs. Starting from a randomly designed library consisting of 9 thio acids and 9 sulfonyl azides leading to 81 potential acylsulfonamides, the target protein, Bcl-X(L), selectively assembled four PPIMs, acylsulfonamides SZ4TA2, SZ7TA2, SZ9TA1, and SZ9TA5, which have been shown to modulate Bcl-X(L)/BH3 interactions. To further investigate the Bcl-X(L) templation effect, control experiments were carried out using two mutants of Bcl-X(L). In one mutant, phenylalanine Phe131 and aspartic acid Asp133, which are critical for the BH3 domain binding, were substituted by alanines, while arginine Arg139, a residue identified to play a crucial role in the binding of ABT-737, a BH3 mimetic, was replaced by an alanine in the other mutant. Incubation of these mutants with the reactive fragments and subsequent LC/MS-SIM analysis confirmed that these building block combinations yield the corresponding acylsulfonamides at the BH3 binding site, the actual "hot spot" of Bcl-X(L). These results validate kinetic TGS using the sulfo-click reaction as a valuable tool for the straightforward identification of high-quality PPIMs.
authors.affiliation: Department of Chemistry, University of South Florida , CHE 205, 4202 E. Fowler Avenue, Tampa, Florida 33620, United States.
authors.forename: Sameer S
authors.initials: SS
authors.lastname: Kulkarni
authors.forename: Xiangdong
authors.initials: X
authors.lastname: Hu
authors.forename: Kenichiro
authors.initials: K
authors.lastname: Doi
authors.forename: Hong-Gang
authors.initials: HG
authors.lastname: Wang
authors.forename: Roman
authors.initials: R
authors.lastname: Manetsch
mesh_terms: D000409:Alanine; D000595:Amino Acid Sequence; D051017:Apoptosis Regulatory Proteins / Q000235:genetics / Q000378:metabolism; D001120:Arginine; D001224:Aspartic Acid; D000072224:Bcl-2-Like Protein 11; D001665:Binding Sites; D001671:Biochemistry / Q000379:methods; D057930:Click Chemistry / Q000379:methods*; D008024:Ligands; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D057786:Peptidomimetics; D010649:Phenylalanine; D054730:Protein Interaction Domains and Motifs; D011518:Proto-Oncogene Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D054852:Small Molecule Libraries; D013449:Sulfonamides / Q000737:chemistry / Q000378:metabolism; D051020:bcl-X Protein / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D051017:Apoptosis Regulatory Proteins; C474007:Bax protein (53-86); D000072224:Bcl-2-Like Protein 11; D008024:Ligands; D008565:Membrane Proteins; D010446:Peptide Fragments; D057786:Peptidomimetics; D011518:Proto-Oncogene Proteins; D054852:Small Molecule Libraries; D013449:Sulfonamides; D051020:bcl-X Protein; D001224:Aspartic Acid; D010649:Phenylalanine; D001120:Arginine; D000409:Alanine