DNA-encoded chemical libraries yield non-covalent and non-peptidic SARS-CoV-2 main protease inhibitors.

The development of SARS-CoV-2 main protease (M) inhibitors for the treatment of COVID-19 has mostly benefitted from X-ray structures and preexisting knowledge of inhibitors; however, an efficient method to generate M inhibitors, which circumvents such information would be advantageous. As an alternative approach, we show here that DNA-encoded chemistry technology (DEC-Tec) can be used to discover inhibitors of M. An affinity selection of a 4-billion-membered DNA-encoded chemical library (DECL) using M as bait produces novel non-covalent and non-peptide-based small molecule inhibitors of M with low nanomolar K values.

Discovery of potent BET bromodomain 1 stereoselective inhibitors using DNA-encoded chemical library selections.

BRDT, BRD2, BRD3, and BRD4 comprise the bromodomain and extraterminal (BET) subfamily which contain two similar tandem bromodomains (BD1 and BD2). Selective BD1 inhibition phenocopies effects of tandem BET BD inhibition both in cancer models and, as we and others have reported of BRDT, in the testes. To find novel BET BD1 binders, we screened >4.

DNA-encoded chemistry technology yields expedient access to SARS-CoV-2 M inhibitors.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has killed more than 4 million humans globally, but there is no bona fide Food and Drug Administration-approved drug-like molecule to impede the COVID-19 pandemic. The sluggish pace of traditional therapeutic discovery is poorly suited to producing targeted treatments against rapidly evolving viruses. Here, we used an affinity-based screen of 4 billion DNA-encoded molecules en masse to identify a potent class of virus-specific inhibitors of the SARS-CoV-2 main protease (M) without extensive and time-consuming medicinal chemistry.

Discovery and characterization of bromodomain 2-specific inhibitors of BRDT.

Bromodomain testis (BRDT), a member of the bromodomain and extraterminal (BET) subfamily that includes the cancer targets BRD2, BRD3, and BRD4, is a validated contraceptive target. All BET subfamily members have two tandem bromodomains (BD1 and BD2). Knockout mice lacking BRDT-BD1 or both bromodomains are infertile.

Discovery of potent thrombin inhibitors from a protease-focused DNA-encoded chemical library.

DNA-encoded chemical libraries are collections of compounds individually coupled to unique DNA tags serving as amplifiable identification barcodes. By bridging split-and-pool combinatorial synthesis with the ligation of unique encoding DNA oligomers, million- to billion-member libraries can be synthesized for use in hundreds of healthcare target screens. Although structural diversity and desirable molecular property ranges generally guide DNA-encoded chemical library design, recent reports have highlighted the utility of focused DNA-encoded chemical libraries that are structurally biased for a class of protein targets.

Identifying Oxacillinase-48 Carbapenemase Inhibitors Using DNA-Encoded Chemical Libraries.

Bacterial resistance to β-lactam antibiotics is largely mediated by β-lactamases, which catalyze the hydrolysis of these drugs and continue to emerge in response to antibiotic use. β-Lactamases that hydrolyze the last resort carbapenem class of β-lactam antibiotics (carbapenemases) are a growing global health threat. Inhibitors have been developed to prevent β-lactamase-mediated hydrolysis and restore the efficacy of these antibiotics.

C-N Coupling of DNA-Conjugated (Hetero)aryl Bromides and Chlorides for DNA-Encoded Chemical Library Synthesis.

DNA-encoded chemical library (DECL) screens are a rapid and economical tool to identify chemical starting points for drug discovery. As a robust transformation for drug discovery, palladium-catalyzed C-N coupling is a valuable synthetic method for the construction of DECL chemical matter; however, currently disclosed methods have only been demonstrated on DNA-attached (hetero)aromatic iodide and bromide electrophiles. We developed conditions utilizing an -heterocyclic carbene-palladium catalyst that extends this reaction to the coupling of DNA-conjugated (hetero)aromatic chlorides with (hetero)aromatic and select aliphatic amine nucleophiles.

Trapping intermediates in metal transfer reactions of the CusCBAF export pump of .

CusCBAF represents an important class of bacterial efflux pump exhibiting selectivity towards Cu(I) and Ag(I). The complex is comprised of three proteins: the CusA transmembrane pump, the CusB soluble adaptor protein, and the CusC outer-membrane pore, and additionally requires the periplasmic metallochaperone CusF. Here we used spectroscopic and kinetic tools to probe the mechanism of copper transfer between CusF and CusB using selenomethionine labeling of the metal-binding Met residues coupled to RFQ-XAS at the Se and Cu edges.

Effects of Active Site Mutations on Specificity of Nucleobase Binding in Human DNA Polymerase η.

Human DNA polymerase η (Pol η) plays a vital role in protection against skin cancer caused by damage from ultraviolet light. This enzyme rescues stalled replication forks at cyclobutane thymine-thymine dimers (TTDs) by inserting nucleotides opposite these DNA lesions. Residue R61 is conserved in the Pol η enzymes across species, but the corresponding residue, as well as its neighbor S62, is different in other Y-family polymerases, Pol ι and Pol κ.

Molecular Dynamics Study of Twister Ribozyme: Role of Mg(2+) Ions and the Hydrogen-Bonding Network in the Active Site.

The recently discovered twister ribozyme is thought to utilize general acid-base catalysis in its self-cleavage mechanism, but the roles of nucleobases and metal ions in the mechanism are unclear. Herein, molecular dynamics simulations of the env22 twister ribozyme are performed to elucidate the structural and equilibrium dynamical properties, as well as to examine the role of Mg(2+) ions and possible candidates for the general base and acid in the self-cleavage mechanism. The active site region and the ends of the pseudoknots were found to be less mobile than other regions of the ribozyme, most likely providing structural stability and possibly facilitating catalysis.

Comparative Molecular Dynamics Studies of Human DNA Polymerase η.

High-energy ultraviolet radiation damages DNA through the formation of cyclobutane pyrimidine dimers, which stall replication. When the lesion is a thymine-thymine dimer (TTD), human DNA polymerase η (Pol η) assists in resuming the replication process by inserting nucleotides opposite the damaged site. We performed extensive molecular dynamics (MD) simulations to investigate the structural and dynamical effects of four different Pol η complexes with or without a TTD and with either dATP or dGTP as the incoming base.

Relative Binding Free Energies of Adenine and Guanine to Damaged and Undamaged DNA in Human DNA Polymerase η: Clues for Fidelity and Overall Efficiency.

Human DNA polymerase η (Pol η) plays an essential protective role against skin cancer caused by cyclobutane thymine-thymine dimers (TTDs), a frequent form of DNA damage arising from exposure to the sun. This enzyme rescues stalled replication forks at the TTDs by inserting bases opposite these DNA defects. Herein we calculate binding free energies for a free deoxyribose nucleotide triphosphate, dATP or dGTP, to Pol η complexed with undamaged or damaged DNA.

Models for the Metal Transfer Complex of the N-Terminal Region of CusB and CusF.

The tripartite CusCFBA pump in Escherichia coli is a very effective heavy metal extrusion system specific for Cu(I) and Ag(I). The N-terminal region of the membrane fusion protein CusB (CusB-NT) is highly disordered, and hence, experimentally characterizing its structure is challenging. In a previous study, this disorder was confirmed with molecular dynamics simulations, although some key structural elements were determined.

Molecular Dynamics Study of Urease.

have been implicated in an array of gastrointestinal disorders including, but not limited to, gastric and duodenal ulcers and adenocarcinoma. This bacterium utilizes an enzyme, urease, to produce copious amounts of ammonia through urea hydrolysis in order to survive the harsh acidic conditions of the stomach. Molecular dynamics (MD) studies on the urease enzyme have been employed in order to study structural features of this enzyme that may shed light on the hydrolysis mechanism.

Bringing Clarity to the Prediction of Protein-Ligand Binding Free Energies via "Blurring".

We present a method to evaluate the free energies of ligand binding utilizing a Monte Carlo estimation of the configuration integrals concomitant with uncertainty quantification. Ensembles for integration are built through systematically perturbing an initial ligand conformation in a rigid binding pocket, which is optimized separately prior to incorporation of the ligand. We call the procedure producing the ensembles "blurring", and it is carried out using an in-house developed code.

The Movable Type Method Applied to Protein-Ligand Binding.

Accurately computing the free energy for biological processes like protein folding or protein-ligand association remains a challenging problem. Both describing the complex intermolecular forces involved and sampling the requisite configuration space make understanding these processes innately difficult. Herein, we address the sampling problem using a novel methodology we term "movable type".

Structure and dynamics of the N-terminal domain of the Cu(I) binding protein CusB.

CusCFBA is one of the metal efflux systems in Escherichia coli that is highly specific for its substrates, Cu(I) and Ag(I). It serves to protect the bacteria in environments that have lethal concentrations of these metals. The membrane fusion protein CusB is the periplasmic piece of CusCFBA, which has not been fully characterized by crystallography because of its extremely disordered N-terminal region.

Prediction of trypsin/molecular fragment binding affinities by free energy decomposition and empirical scores.

Two families of binding affinity estimation methodologies are described which were utilized in the SAMPL3 trypsin/fragment binding affinity challenge. The first is a free energy decomposition scheme based on a thermodynamic cycle, which included separate contributions from enthalpy and entropy of binding as well as a solvent contribution. Enthalpic contributions were estimated with PM6-DH2 semiempirical quantum mechanical interaction energies, which were modified with a statistical error correction procedure.

Insights into the mechanistic dichotomy of the protein farnesyltransferase peptide substrates CVIM and CVLS.

Protein farnesyltransferase (FTase) catalyzes farnesylation of a variety of peptide substrates. (3)H α-secondary kinetic isotope effect (α-SKIE) measurements of two peptide substrates, CVIM and CVLS, are significantly different and have been proposed to reflect a rate-limiting S(N)2-like transition state with dissociative characteristics for CVIM, while, due to the absence of an isotope effect, CVLS was proposed to have a rate-limiting peptide conformational change. Potential of mean force quantum mechanical/molecular mechanical studies coupled with umbrella sampling techniques were performed to further probe this mechanistic dichotomy.

Insight into the cation-π interaction at the metal binding site of the copper metallochaperone CusF.

The periplasmic Cu(+)/Ag(+) chaperone CusF features a novel cation-π interaction between a Cu(+)/Ag(+) ion and Trp44 at the metal binding site. The nature and strength of the Cu(+)/Ag(+)-Trp44 interactions were investigated using computational methodologies. Quantum-mechanical (QM) calculations showed that the Cu(+) and Ag(+) interactions with Trp44 are of similar strength (~14 kcal/mol) and bond order.

Origins of the diastereoselectivity in hydrogen bonding directed Diels-Alder reactions of chiral dienes with achiral dienophiles: a computational study.

In this study, the origins of diastereoselectivity in the hydrogen bonding assisted Diels-Alder reactions of chiral dienes with achiral dienophiles have been investigated with density functional methods. The distortion/interaction model has been applied to shed light on the origins of selectivity. C9-Substituted chiral anthracene templates (R = (CH(3))(OCH(3))(H), R = (CH(3))(OH)(H), R = (CH(3))(CH(2)CH(3))(H) and R = (-CH(2)-C(CH(3))(OCH(3))(H)) are used to rationalize the role of a stereogenic center and H-bonding on the product distribution ratio.

Pairwise additivity of energy components in protein-ligand binding: the HIV II protease-Indinavir case.

An energy expansion (binding energy decomposition into n-body interaction terms for n ≥ 2) to express the receptor-ligand binding energy for the fragmented HIV II protease-Indinavir system is described to address the role of cooperativity in ligand binding. The outcome of this energy expansion is compared to the total receptor-ligand binding energy at the Hartree-Fock, density functional theory, and semiempirical levels of theory. We find that the sum of the pairwise interaction energies approximates the total binding energy to ∼82% for HF and to >95% for both the M06-L density functional and PM6-DH2 semiempirical method.