ABSTRACT
The recognition of methyl-lysine and -arginine residues on both histone and other proteins by specific "reader" elements is important for chromatin regulation, gene expression, and control of cell-cycle progression. Recently the crucial role of these reader proteins in cancer development and dedifferentiation has emerged, owing to the increased interest among the scientific community. The methyl-lysine and -arginine readers are a large and very diverse set of effector proteins and targeting them with small molecule probes in drug discovery will inevitably require a detailed understanding of their structural biology and mechanism of binding. In the following review, the critical elements of methyl-lysine and -arginine recognition will be summarized with respect to each protein family and initial results in assay development, probe design, and drug discovery will be highlighted.
ABSTRACT
Proteins which bind methylated lysines ("readers" of the histone code) are important components in the epigenetic regulation of gene expression and can also modulate other proteins that contain methyl-lysine such as p53 and Rb. Recognition of methyl-lysine marks by MBT domains leads to compaction of chromatin and a repressed transcriptional state. Antagonists of MBT domains would serve as probes to interrogate the functional role of these proteins and initiate the chemical biology of methyl-lysine readers as a target class. Small-molecule MBT antagonists were designed based on the structure of histone peptide-MBT complexes and their interaction with MBT domains determined using a chemiluminescent assay and ITC. The ligands discovered antagonize native histone peptide binding, exhibiting 5-fold stronger binding affinity to L3MBTL1 than its preferred histone peptide. The first cocrystal structure of a small molecule bound to L3MBTL1 was determined and provides new insights into binding requirements for further ligand design.
Subject(s)
Lysine/metabolism , Nuclear Proteins/metabolism , Small Molecule Libraries/metabolism , Binding Sites , Calorimetry , Drug Discovery , High-Throughput Screening Assays , Humans , Ligands , Luminescent Measurements , Methylation , Models, Molecular , Nuclear Proteins/antagonists & inhibitors , Nuclear Proteins/chemistry , Peptidomimetics/chemistry , Peptidomimetics/metabolism , Peptidomimetics/pharmacology , Protein Structure, Tertiary , Small Molecule Libraries/chemistry , Small Molecule Libraries/pharmacologyABSTRACT
Thiol-thioester exchange was found to readily generate libraries of cyclic thiodepsipeptides under thermodynamic control, which will enable their use in a variety of dynamic combinatorial chemistry assays. The kinetic determinants of macrocycle formation and the role of amino acid structure on the reaction dynamics are discussed.
Subject(s)
Combinatorial Chemistry Techniques , Depsipeptides/chemistry , Esters/chemistry , Peptides, Cyclic/chemistry , Sulfhydryl Compounds/chemistry , KineticsABSTRACT
A small molecule receptor that mimics the HP1 chromodomain's affinity for trimethyl lysine has been identified from a dynamic combinatorial library. Discrimination for trimethyl lysine over the lower methylation states parallels that of the native protein receptor. These studies demonstrate the feasibility of small molecule receptors as potential sensors for protein post-translational modifications.
Subject(s)
Histones/chemistry , Lysine/chemistry , Amino Acid Substitution , Combinatorial Chemistry Techniques , Histones/genetics , Methylation , Mutagenesis, Site-Directed , Protein Processing, Post-TranslationalABSTRACT
The novel azobenzene-based monomer 1 was prepared, equipped with the necessary functionality to undergo simultaneous dynamic exchange processes: hydrazone exchange and photoisomerization. Acid-promoted hydrolysis of the azobenzene building block produced a dynamic combinatorial library of cyclic oligomers, while multibuilding block libraries were also generated upon addition of proline-based monomers. Libraries equilibrated under thermal conditions were dominated by trans isomers of the azobenzene macrocycles, whereas light-induced isomerization resulted in a conformational change of the library members to their corresponding cis-azo form. In the presence of a pentaproline template, a slower rate of thermal relaxation of the cis-azobenzene species 1(c) was observed, resulting in stabilization and amplification of this receptor due to favorable binding interactions. The facile identification and application of such photoswitchable receptors have the potential to allow for greater control over molecular recognition events.
