Expanding the druggable space of the LSD1/CoREST epigenetic target: new potential binding regions for drug-like molecules, peptides, protein partners, and chromatin.

Lysine specific demethylase-1 (LSD1/KDM1A) in complex with its corepressor protein CoREST is a promising target for epigenetic drugs. No therapeutic that targets LSD1/CoREST, however, has been reported to date. Recently, extended molecular dynamics (MD) simulations indicated that LSD1/CoREST nanoscale clamp dynamics is regulated by substrate binding and highlighted key hinge points of this large-scale motion as well as the relevance of local residue dynamics. Prompted by the urgent need for new molecular probes and inhibitors to understand LSD1/CoREST interactions with small-molecules, peptides, protein partners, and chromatin, we undertake here a configurational ensemble approach to expand LSD1/CoREST druggability. The independent algorithms FTMap and SiteMap and our newly developed Druggable Site Visualizer (DSV) software tool were used to predict and inspect favorable binding sites. We find that the hinge points revealed by MD simulations at the SANT2/Tower interface, at the SWIRM/AOD interface, and at the AOD/Tower interface are new targets for the discovery of molecular probes to block association of LSD1/CoREST with chromatin or protein partners. A fourth region was also predicted from simulated configurational ensembles and was experimentally validated to have strong binding propensity. The observation that this prediction would be prevented when using only the X-ray structures available (including the X-ray structure bound to the same peptide) underscores the relevance of protein dynamics in protein interactions. A fifth region was highlighted corresponding to a small pocket on the AOD domain. This study sets the basis for future virtual screening campaigns targeting the five novel regions reported herein and for the design of LSD1/CoREST mutants to probe LSD1/CoREST binding with chromatin and various protein partners

Protein Recognition by Short Peptide Reversible Inhibitors of the Chromatin-Modifying LSD1/CoREST Lysine Demethylase.

The combinatorial assembly of protein complexes is at the heart of chromatin biology. Lysine demethylase LSD1(KDM1A)/CoREST beautifully exemplifies this concept. The active site of the enzyme tightly associates to the N-terminal domain of transcription factors of the SNAIL1 family, which therefore can competitively inhibit the binding of the N-terminal tail of the histone substrate. Our enzymatic, crystallographic, spectroscopic, and computational studies reveal that LSD1/CoREST can bind to a hexapeptide derived from the SNAIL sequence through recognition of a positively charged α-helical turn that forms upon binding to the enzyme. Variations in sequence and length of this six amino acid ligand modulate affinities enabling the same binding site to differentially interact with proteins that exert distinct biological functions. The discovered short peptide inhibitors exhibit antiproliferative activities and lay the foundation for the development of peptidomimetic small molecule inhibitors of LSD1

Protein Recognition by Short Peptide Reversible Inhibitors of the Chromatin-Modifying LSD1/CoREST Lysine Demethylase

The combinatorial assembly of protein complexes is at the heart of chromatin biology. Lysine demethylaseLSD1(KDM1A)/CoREST beautifully exemplifies this concept. The active site of the enzyme tightly associates to the N-terminaldomain of transcription factors of the SNAIL1 family, which therefore can competitively inhibit the binding of the N-terminal tailof the histone substrate. Our enzymatic, crystallographic, spectroscopic, and computational studies reveal that LSD1/CoRESTcan bind to a hexapeptide derived from the SNAIL sequence through recognition of a positively chargedα-helical turn that formsupon binding to the enzyme. Variations in sequence and length of this six amino acid ligand modulate affinities enabling the samebinding site to differentially interact with proteins that exert distinct biological functions. The discovered short peptide inhibitorsexhibit antiproliferative activities and lay the foundation for the development of peptidomimetic small molecule inhibitors ofLSD

Interplay among nucleosomal DNA, histone tails, and corepressor CoREST underlies LSD1-mediated H3 demethylation.

With its noncatalytic domains, DNA-binding regions, and a catalytic core targeting the histone tails, LSD1-CoREST (lysine-specific demethylase 1; REST corepressor) is an ideal model system to study the interplay between DNA binding and histone modification in nucleosome recognition. To this end, we covalently associated LSD1-CoREST to semisynthetic nucleosomal particles. This enabled biochemical and biophysical characterizations of nucleosome binding and structural elucidation by small-angle X-ray scattering, which was extensively validated through binding assays and site-directed mutagenesis of functional interfaces. Our results suggest that LSD1-CoREST functions as an ergonomic clamp that induces the detachment of the H3 histone tail from the nucleosomal DNA to make it available for capture by the enzyme active site. The key notion emerging from these studies is the inherently competitive nature of the binding interactions because nucleosome tails, chromatin modifiers, transcription factors, and DNA represent sites for multiple and often mutually exclusive interactions

Molecular Insights into Human Monoamine Oxidase B Inhibition by the Glitazone Antidiabetes Drugs

The widely employed antidiabetic drug pioglitazone (Actos) is shown to be a specific and reversible inhibitor of human monoamine oxidase B (MAO B). The crystal structure of the enzyme–inhibitor complex shows that the <i>R</i>-enantiomer is bound with the thiazolidinedione ring near the flavin. The molecule occupies both substrate and entrance cavities of the active site, establishing noncovalent interactions with the surrounding amino acids. These binding properties differentiate pioglitazone from the clinically used MAO inhibitors, which act through covalent inhibition mechanisms and do not exhibit a high degree of MAO A versus B selectivity. Rosiglitazone (Avandia) and troglitazone, other members of the glitazone class, are less selective in that they are weaker inhibitors of both MAO A and MAO B. These results suggest that pioglitazone may have utility as a “repurposed” neuroprotectant drug in retarding the progression of disease in Parkinson's patients. They also provide new insights for the development of reversible isoenzyme-specific MAO inhibitors

Interplay among nucleosomal DNA, histone tails, and corepressor CoREST underlies LSD1-mediated H3 demethylation.

International audienceWith its noncatalytic domains, DNA-binding regions, and a catalytic core targeting the histone tails, LSD1-CoREST (lysine-specific demethylase 1; REST corepressor) is an ideal model system to study the interplay between DNA binding and histone modification in nucleosome recognition. To this end, we covalently associated LSD1-CoREST to semisynthetic nucleosomal particles. This enabled biochemical and biophysical characterizations of nucleosome binding and structural elucidation by small-angle X-ray scattering, which was extensively validated through binding assays and site-directed mutagenesis of functional interfaces. Our results suggest that LSD1-CoREST functions as an ergonomic clamp that induces the detachment of the H3 histone tail from the nucleosomal DNA to make it available for capture by the enzyme active site. The key notion emerging from these studies is the inherently competitive nature of the binding interactions because nucleosome tails, chromatin modifiers, transcription factors, and DNA represent sites for multiple and often mutually exclusive interactions

Interplay among nucleosomal DNA, histone tails, and corepressor CoREST underlies LSD1-mediated H3 demethylation.

International audienceWith its noncatalytic domains, DNA-binding regions, and a catalytic core targeting the histone tails, LSD1-CoREST (lysine-specific demethylase 1; REST corepressor) is an ideal model system to study the interplay between DNA binding and histone modification in nucleosome recognition. To this end, we covalently associated LSD1-CoREST to semisynthetic nucleosomal particles. This enabled biochemical and biophysical characterizations of nucleosome binding and structural elucidation by small-angle X-ray scattering, which was extensively validated through binding assays and site-directed mutagenesis of functional interfaces. Our results suggest that LSD1-CoREST functions as an ergonomic clamp that induces the detachment of the H3 histone tail from the nucleosomal DNA to make it available for capture by the enzyme active site. The key notion emerging from these studies is the inherently competitive nature of the binding interactions because nucleosome tails, chromatin modifiers, transcription factors, and DNA represent sites for multiple and often mutually exclusive interactions