Inhibition of Sirtuin Deacylase Activity by Peroxynitrite.

Sirtuins are a class of enzymes that deacylate protein lysine residues using NAD as a cosubstrate. Sirtuin deacylase activity has been historically regarded as protective; loss of sirtuin deacylase activity potentially increases susceptibility to aging-related disease development. However, which factors may inhibit sirtuins during aging or disease is largely unknown.

Oxidized LDL regulates efferocytosis through the CD36-PKM2-mtROS pathway.

Unlabelled: Macrophage efferocytosis, the process by which phagocytes engulf and remove apoptotic cells (ACs), plays a critical role in maintaining tissue homeostasis. Efficient efferocytosis prevents secondary necrosis, mitigates chronic inflammation, and impedes atherosclerosis progression. However, the regulatory mechanisms of efferocytosis under atherogenic conditions remain poorly understood.

Zinc-chelating BET bromodomain inhibitors equally target islet endocrine cell types.

Inhibition of the bromodomain and extraterminal domain (BET) protein family is a potential strategy to prevent and treat diabetes; however, the clinical use of BET bromodomain inhibitors (BETis) is associated with adverse effects. Here, we explore a strategy for targeting BETis to β cells by exploiting the high-zinc (Zn) concentration in β cells relative to other cell types. We report the synthesis of a novel, Zn-chelating derivative of the pan-BETi (+)-JQ1, (+)-JQ1-DPA, in which (+)-JQ1 was conjugated to dipicolyl amine (DPA).

Current Trends in Sirtuin Activator and Inhibitor Development.

Sirtuins are NAD-dependent protein deacylases and key metabolic regulators, coupling the cellular energy state with selective lysine deacylation to regulate many downstream cellular processes. Humans encode seven sirtuin isoforms (Sirt1-7) with diverse subcellular localization and deacylase targets. Sirtuins are considered protective anti-aging proteins since increased sirtuin activity is canonically associated with lifespan extension and decreased activity with developing aging-related diseases.

Cancer-associated polybromo-1 bromodomain 4 missense variants variably impact bromodomain ligand binding and cell growth suppression.

The polybromo, brahma-related gene 1-associated factors (PBAF) chromatin remodeling complex subunit polybromo-1 (PBRM1) contains six bromodomains that recognize and bind acetylated lysine residues on histone tails and other nuclear proteins. PBRM1 bromodomains thus provide a link between epigenetic posttranslational modifications and PBAF modulation of chromatin accessibility and transcription. As a putative tumor suppressor in several cancers, PBRM1 protein expression is often abrogated by truncations and deletions.

Extrahelical Binding Site for a 1-Imidazo[4,5-c]quinolin-4-amine A Adenosine Receptor Positive Allosteric Modulator on Helix 8 and Distal Portions of Transmembrane Domains 1 and 7.

This study describes the localization and computational prediction of a binding site for the A adenosine receptor (AAR) positive allosteric modulator 2-cyclohexyl-1-imidazo[4,5-c]quinolin-4-(3,4-dichlorophenyl)amine (LUF6000). The work reveals an extrahelical lipid-facing binding pocket disparate from the orthosteric binding site that encompasses transmembrane domain (TMD) 1, TMD7, and Helix (H) 8, which was predicted by molecular modeling and validated by mutagenesis. According to the model, the nearly planar 1-imidazo[4,5-c]quinolinamine ring system lies parallel to the transmembrane segments, inserted into an aromatic cage formed by π-π stacking interactions with the side chains of Y284 in TMD7 and Y293 in H8 and by π-NH bonding between Y284 and the exocyclic amine.

Identification of Protein Targets of -Nitroso-Coenzyme A-Mediated -Nitrosation Using Chemoproteomics.

-Nitrosation is a cysteine post-translational modification fundamental to cellular signaling. This modification regulates protein function in numerous biological processes in the nervous, cardiovascular, and immune systems. Small molecule or protein nitrosothiols act as mediators of NO signaling by transferring the NO group (formally NO) to a free thiol on a target protein through a transnitrosation reaction.

Deep computational phenotyping of genomic variants impacting the SET domain of KMT2C reveal molecular mechanisms for their dysfunction.

Kleefstra Syndrome type 2 (KLEFS-2) is a genetic, neurodevelopmental disorder characterized by intellectual disability, infantile hypotonia, severe expressive language delay, and characteristic facial appearance, with a spectrum of other distinct clinical manifestations. Pathogenic mutations in the epigenetic modifier type 2 lysine methyltransferase KMT2C have been identified to be causative in KLEFS-2 individuals. This work reports a translational genomic study that applies a multidimensional computational approach for deep variant phenotyping, combining conventional genomic analyses, advanced protein bioinformatics, computational biophysics, biochemistry, and biostatistics-based modeling.

A multi-layered computational structural genomics approach enhances domain-specific interpretation of Kleefstra syndrome variants in EHMT1.

This study investigates the functional significance of assorted variants of uncertain significance (VUS) in euchromatic histone lysine methyltransferase 1 (EHMT1), which is critical for early development and normal physiology. EHMT1 mutations cause Kleefstra syndrome and are linked to various human cancers. However, accurate functional interpretations of these variants are yet to be made, limiting diagnoses and future research.

Fragment-based screening by protein-detected NMR spectroscopy.

Fragment-based drug discovery (FBDD) identifies low molecular weight compounds that can be developed into ligands with high affinity and selectivity for therapeutic targets. Screening fragment libraries (<10,000 molecules) with biophysical techniques against macromolecules provides information about novel chemical spaces that bind the macromolecule and scaffolds that can be modified to increase potency. A fragment-screening pipeline requires a standardized protocol for target selection, library assembly and maintenance, library screening, and hit validation to ensure hit integrity.

Analysis of continuous enzyme kinetic data using ICEKAT.

ICEKAT (Interactive Continuous Enzyme Analysis Tool) is an interactive web-based program for calculating initial rates and kinetic parameters (e.g., V, k, K, EC, IC) from continuous enzyme kinetic assay data that satisfy Michaelis-Menten and steady-state kinetic assumptions.

Beyond structural bioinformatics for genomics with dynamics characterization of an expanded KRAS mutational landscape.

Current capabilities in genomic sequencing outpace functional interpretations. Our previous work showed that 3D protein structure calculations enhance mechanistic understanding of genetic variation in sequenced tumors and patients with rare diseases. The KRAS GTPase is among the critical genetic factors driving cancer and germline conditions.

A Multi-Layered Computational Structural Genomics Approach Enhances Domain-Specific Interpretation of Kleefstra Syndrome Variants in EHMT1.

This study investigates the functional significance of assorted variants of uncertain significance (VUS) in euchromatic histone lysine methyltransferase 1 (EHMT1), which is critical for early development and normal physiology. EHMT1 mutations cause Kleefstra syndrome and are linked to various human cancers. However, accurate functional interpretation of these variants are yet to be made, limiting diagnoses and future research.

Rational Design and Development of Selective BRD7 Bromodomain Inhibitors and Their Activity in Prostate Cancer.

Bromodomain-containing proteins are readers of acetylated lysine and play important roles in cancer. Bromodomain-containing protein 7 (BRD7) is implicated in multiple malignancies; however, there are no selective chemical probes to study its function in disease. Using crystal structures of BRD7 and BRD9 bromodomains (BDs) bound to BRD9-selective ligands, we identified a binding pocket exclusive to BRD7.

Synthesis of fluorinated triphenylphosphonium analogs that improve cancer cell selectivity and in vivo detection.

Triphenylphosphonium (TPP) compounds like mito-metformin (MMe) target cancer cells by exploiting their hyperpolarized mitochondrial membrane potential. Here, we present a protocol for synthesizing TPP analogs with selectivity for mammalian cancer cells, reduced toxicity, and quantifiability using fluorine-19 nuclear magnetic resonance (F-NMR). We describe steps for treating mammalian cells with mitochondria-targeted compounds, treating and preparing mouse tissue with these compounds, and F-NMR detection of MMe analogs in cells and tissue.

Cysteine and methionine oxidation in thrombotic disorders.

Thrombosis is the leading cause of death in many diseased conditions. Oxidative stress is characteristic of these conditions. Yet, the mechanisms through which oxidants become prothrombotic are unclear.

Beyond Structural Bioinformatics for Genomics with Dynamics Characterization of an Expanded KRAS Mutational Landscape.

Current capabilities in genomic sequencing outpace functional interpretations. Our previous work showed that 3D protein structure calculations enhance mechanistic understanding of genetic variation in sequenced tumors and patients with rare diseases. The KRAS GTPase is among the critical genetic factors driving cancer and germline conditions.

Sulfenylation links oxidative stress to protein disulfide isomerase oxidase activity and thrombus formation.

Background: Oxidative stress contributes to thrombosis in atherosclerosis, inflammation, infection, aging, and malignancy. Oxidant-induced cysteine modifications, including sulfenylation, can act as a redox-sensitive switch that controls protein function. Protein disulfide isomerase (PDI) is a prothrombotic enzyme with exquisitely redox-sensitive active-site cysteines.

Harnessing the Noncanonical Keap1-Nrf2 Pathway for Human Cytomegalovirus Control.

Host-derived cellular pathways can provide an unfavorable environment for virus replication. These pathways have been a subject of interest for herpesviruses, including the betaherpesvirus human cytomegalovirus (HCMV). Here, we demonstrate that a compound, ARP101, induces the noncanonical sequestosome 1 (SQSTM1)/p62-Keap1-Nrf2 pathway for HCMV suppression.

Fragment-based drug discovery of small molecule ligands for the human chemokine CCL28.

The mucosal chemokine CCL28 is a promising target for immunotherapy drug development due to its elevated expression level in epithelial cells and critical role in creating and maintaining an immunosuppressive tumor microenvironment. Using sulfotyrosine as a probe, NMR chemical shift mapping identified a potential receptor-binding hotspot on the human CCL28 surface. CCL28 was screened against 2,678 commercially available chemical fragments by 2D NMR, yielding thirteen verified hits.

Fluorinated triphenylphosphonium analogs improve cell selectivity and detection of mito-metformin.

Triphenylphosphonium (TPP) conjugated compounds selectively target cancer cells by exploiting their hyperpolarized mitochondrial membrane potential. To date, studies have focused on modifying either the linker or the cargo of TPP-conjugated compounds. Here, we investigated the biological effects of direct modification to TPP to improve the efficacy and detection of mito-metformin (MMe), a TPP-conjugated probe we have shown to have promising preclinical efficacy against solid cancer cells.

Pharmacological inhibition of the acetyltransferase Tip60 mitigates myocardial infarction injury.

Pharmacologic strategies that target factors with both pro-apoptotic and anti-proliferative functions in cardiomyocytes (CMs) may be useful for the treatment of ischemic heart disease. One such multifunctional candidate for drug targeting is the acetyltransferase Tip60, which is known to acetylate both histone and non-histone protein targets that have been shown in cancer cells to promote apoptosis and to initiate the DNA damage response, thereby limiting cellular expansion. Using a murine model, we recently published findings demonstrating that CM-specific disruption of the Kat5 gene encoding Tip60 markedly protects against the damaging effects of myocardial infarction (MI).