ABSTRACT
Despite unequivocal roles in disease, transcription factors (TFs) remain largely untapped as pharmacologic targets due to the challenges in targeting protein-protein and protein-DNA interactions. Here we report a chemical strategy to generate modular synthetic transcriptional repressors (STRs) derived from the bHLH domain of MAX. Our synthetic approach yields chemically stabilized tertiary domain mimetics that cooperatively bind the MYC/MAX consensus E-box motif with nanomolar affinity, exhibit specificity that is equivalent to or beyond that of full-length TFs and directly compete with MYC/MAX protein for DNA binding. A lead STR directly inhibits MYC binding in cells, downregulates MYC-dependent expression programs at the proteome level and inhibits MYC-dependent cell proliferation. Co-crystallization and structure determination of a STR:E-box DNA complex confirms retention of DNA recognition in a near identical manner as full-length bHLH TFs. We additionally demonstrate structure-blind design of STRs derived from alternative bHLH-TFs, confirming that STRs can be used to develop highly specific mimetics of TFs targeting other gene regulatory elements.
Subject(s)
Proto-Oncogene Proteins c-myc , Transcription Factors , Proto-Oncogene Proteins c-myc/genetics , Proto-Oncogene Proteins c-myc/chemistry , Proto-Oncogene Proteins c-myc/metabolism , Transcription Factors/genetics , Basic-Leucine Zipper Transcription Factors/genetics , Basic-Leucine Zipper Transcription Factors/metabolism , Helix-Loop-Helix Motifs , Regulatory Sequences, Nucleic Acid , DNA/genetics , DNA/metabolismABSTRACT
Macrocyclization can improve bioactive peptide ligands through preorganization of molecular topology, leading to improvement of pharmacologic properties like binding affinity, cell permeability, and metabolic stability. Here we demonstrate that Diels-Alder [4 + 2] cycloadditions can be harnessed for peptide macrocyclization and stabilization within a range of peptide scaffolds and chemical environments. Diels-Alder cyclization of diverse diene-dienophile reactive pairs proceeds rapidly, in high yield and with tunable stereochemical preferences on solid-phase or in aqueous solution. This reaction can be applied alone or in concert with other stabilization chemistries, such as ring-closing olefin metathesis, to stabilize loop, turn, and α-helical secondary structural motifs. NMR and molecular dynamics studies of model loop peptides confirmed preferential formation of endo cycloadduct stereochemistry, imparting significant structural rigidity to the peptide backbone that resulted in augmented protease resistance and increased biological activity of a Diels-Alder cyclized (DAC) RGD peptide. Separately, we demonstrated the stabilization of DAC α-helical peptides derived from the ERα-binding protein SRC2. We solved a 2.25 Å cocrystal structure of one DAC helical peptide bound to ERα, which unequivocally corroborated endo stereochemistry of the resulting Diels-Alder adduct, and confirmed that the unique architecture of stabilizing motifs formed with this chemistry can directly contribute to target binding. These data establish Diels-Alder cyclization as a versatile approach to stabilize diverse protein structural motifs under a range of chemical environments.
Subject(s)
Cycloaddition Reaction , Macrocyclic Compounds/chemistry , Peptides/chemistry , Models, Molecular , Molecular Structure , Peptide Hydrolases/metabolism , Protein ConformationABSTRACT
An efficient and mild synthesis of 9H-fluorene derivatives through a Pd(0)-catalyzed cross-coupling reaction of 1,1-diboronates with 2,2'-dibromobiphenyls has been developed. This reaction features high yields, operational simplicity, and mild reaction conditions, thus providing an excellent alternative to published methods for 9H-fluorene synthesis.
Subject(s)
Biphenyl Compounds/chemistry , Boronic Acids/chemistry , Fluorenes/chemical synthesis , Catalysis , Fluorenes/chemistry , Molecular Structure , Palladium/chemistryABSTRACT
Palladium-catalyzed cross-coupling reactions of 1,1-diboronates with vinyl bromides and dibromoalkenes were found to afford 1,4-dienes and allenes, respectively. These reactions utilize the high reactivities of both 1,1-diboronates and allylboron intermediates generated in the initial coupling.
ABSTRACT
A convenient method is developed to synthesize 1,1-diboronates from the corresponding N-tosylhydrazones. This method is also applicable to synthesize 1-silyl-1-boron compounds. Meanwhile, derivatization and consecutive Pd-catalyzed cross-coupling reactions with 1,1-boronates were explored, demonstrating the synthetic potential of 1,1-diboronates.
