Triphosphate Binding Research Articles

Comparative Transcriptome and Metabolome Analyses Revealed Quality Difference between Beauty Tea Processed through Indoor Withering and Outdoor Solar Withering.

Withering is the first processing procedure of beauty tea, and there are few reports on the impact of withering methods on the quality of beauty tea and its regulatory mechanisms. Through comparison of fresh tea leaves (FT) with the leaves at Indoor natural Withering 18h (IWT-18) and Outdoor Solar Withering 6h (OWT-6), which were collected at the end of the two withering processes, 17282 and 13984 differentially expressed genes (DEGs) were screened and 267 and 154 differential metabolites (DMs) were identified, respectively. Coexpression network revealed that a large number of DEGs and DMs were enriched in phenylpropanoid, flavonoid, and adenosine Triphosphate binding (ABC) transporter pathways, and the number of DMs and DEGs in IWT-18 vs FT exceeded that in OWT-6 vs FT. Both withering methods promoted a significant increase in content of phenylalanine and upregulation of β-glucoside (BGLU) expression in the phenylpropanoid metabolism pathway. The five proanthocyanidins related to theaflavins in the flavonoid synthesis pathway was more significant accumulation in FT vs IWT-18 than in FT vs OWT-6. Meanwhile, both withering methods can affect the ABC transporter pathway to promote the accumulation of amino acids and their derivatives, but different withering methods affect different ABC transporter families. Outdoor withering with more severe abiotic stress has a greater impact on the ABCG family, while indoor withering has a more significant effect on the ABCC family. Sensory evaluation results showed that the dry tea of IWT-18 was slightly better than that of OWT-6 because of longer withering time and more thorough substance transformation. In conclusion, the formation of honey flavor in beauty tea may be closely related to the DEGs and DMs in these three pathways. Our research provides theoretical data support for further revealing the mechanism of quality formation during the withering process of beauty tea. This article is protected by copyright. All rights reserved.

301 Measuring ATPase activity of adenosine triphosphate–binding cassette subfamily C member 4, lamprey CFTR, and human CFTR using antimony-phosphomolybdate assay

301 Measuring ATPase activity of adenosine triphosphate–binding cassette subfamily C member 4, lamprey CFTR, and human CFTR using antimony-phosphomolybdate assay

Recurrent Harlequin Ichthyosis in a Family: A Case Report

Introduction: Harlequin ichthyosis is a severe form of erythrodermic ichthyosis that manifests in a distinctive and distressing appearance at birth. The newborn baby is covered in thick, plate-like scales with distorted surface features, resulting in limited mobility. Fortunately, this condition is rare, with an estimated incidence of approximately 1 in 300,000 births. It is inherited in an autosomal recessive manner and is caused by a mutation in the adenosine triphosphate binding cassette subfamily A member 12 (ABCA12) gene. This mutation affects lipid transport, and the activity of ABCA12 is crucial to produce long-chain ceramides, which are essential for the development of a normal skin barrier. Methods: We present a case report of a newborn who presented with thick, firmly adherent dense plaques covering the entire body, accompanied by deep fissures that extended into the dermis. The facial features included severe ectropion, conjunctival chemosis, eclabium, tethered ears, and a rudimentary nose. In addition, there was flexion contracture of the arms, legs, and digits, along with hypoplastic nails. Results: The baby experienced respiratory difficulty, poor sucking, and a severe cutaneous infection. Unfortunately, the baby’s condition deteriorated and passed away on the 20th day after delivery. Conclusion: Previously considered universally fatal, harlequin ichthyosis now has reported cases of survivors, although it remains a challenging condition. It is crucial to provide parents and family with comprehensive information about the implications of the condition. They should be encouraged to explore all available options and receive psychological support throughout the process. It is essential for the neonatal intensive care unit staff to be trained in providing specialized care for these babies. In addition, parents should be offered the option of antenatal counseling, particularly in cases where there is a previously affected sibling or a history of consanguinity.

Supplementation of chestnut tannins in diets can improve meat quality and antioxidative capability in Hu lambs

Chestnut tannins (CNT), as a source of hydrolyzable tannins, positively affect the antioxidant status of livestock. In the current study, 90 male Hu lambs were used to investigate the effect of dietary CNT intake on growth performance, nutrient digestibility, meat quality and oxidative stability, rumen microbial, and the transcriptomes of muscle and liver. A completely randomized design with three CNT intake levels (0, 0.3%, and 0.6%) was used. Rumen microbial and nutrient digestibility were not significantly altered by CNT intake. Diets with 0.3% CNT intake significantly reduced the shear force, yellowness at 24 h, and C20:2 polyunsaturated fatty acids of lamb meat and malondialdehyde in serum and longissimus thoracis (LT) muscle. Meanwhile, the 0.3% CNT diet significantly increased average daily gain during the 1– 21 days and 64– 90 days, dry matter intake during the 1– 21 days, the slaughter weight, and liver index of lambs. The 0.3% CNT diet significantly increased C26:0 saturated fatty acids, total antioxidant capacity, glutathione peroxidase, superoxide dismutase, and catalase in LT muscle. The meat shelf life of 0.3% CNT and 0.6% CNT groups was prolonged by 8.7 h and 5.4 h, respectively. Transcriptomic analysis revealed that CNT supplementation can induce the expression of antioxidant enzyme gene (CAT, SOD1), and the differentially expressed genes were mainly involved in antioxidant activity, transferase activity, and adenosine triphosphate binding. These results suggest that 0.3% CNT intake can relieve the oxidative stress of lambs, and improve the stability of meat color and meat tenderness, due to the enhanced antioxidative capacity.

Molecular Insights on the Conformational Transitions and Activity Regulation of the c-Met Kinase Induced by Ligand Binding.

The tyrosine-protein kinase Met (c-Met) is an important signaling molecule involved in cellular growth and division. The dysregulation of c-Met may induce many fatal diseases, including non-small cell lung cancer, gastrointestinal cancers, hepatocellular carcinoma, etc. The activation of the c-Met kinase is dominant by the structure and dynamics of many important functional motifs, which are regulated by adenosine triphosphate (ATP) binding. c-Met inhibitors bind to the ATP-binding site or the allosteric pocket to compete with ATP molecules or alter the conformation of the function-related domains. Nevertheless, the mechanisms of ligand binding to c-Met are still unclear, especially the regulation of the functional motifs by different inhibitors. These greatly impede the development of novel drugs to overcome the drug tolerance to the currently marketed c-Met inhibitors. In this study, we used enhanced sampling technology to study the binding and regulation of two specific c-Met inhibitors. The results show that the two ligands adopt different binding processes even though with similar binding affinity. More importantly, our results uncovered different protein conformational features and the correlated motions of functional motifs regulated by the inhibitors, providing the structural basis for the functional suppression of the protein kinases.

Proteomic investigation and understanding on IgY purification and product development

An increasing demand for the development of immunoglobin Y (IgY) illustrates the necessity of the component analysis in the process of conduction and quality control. This study investigated the proteomic changes in crude IgY extracts and purified IgY products obtained by sequential polyethylene glycol precipitation (PEG) of egg yolks followed by human mycoplasma protein-based affinity chromatography compared with intact egg yolks. After confirming the extraction efficiency and purity by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis, liquid chromatography tandem-mass spectrometry (LC-MS/MS) was performed with samples including fresh yolk, IgY extracted product and purified product. A total of 348 proteins were identified, with 36 proteins deleted and 209 newly detected proteins in the purified product compared to the intact egg yolk. The significantly decreased proteins mainly included phosvitin, albumin, and apolipoprotein B whereas the significantly increased proteins were mainly IgY-related proteins. GO analysis showed that the purified IgY product had ATPase activity and purine ribonucleoside triphosphate binding activity, and was mainly involved in purine and nucleic acid metabolism. This study will inevitably fasten the commercial application of IgY antibodies and is of greater significance for promotion, development and approval for new antibody derived drug products.

Structure-guided discovery of adenosine triphosphate-competitive casein kinase 2 inhibitors.

Casein kinase 2 (CK2) is a ubiquitous, highly pleiotropic serine-threonine kinase. CK2 has been identified as a potential drug target for the treatment of cancer and related disorders. Several adenosine triphosphate-competitive CK2 inhibitors have been identified and have progressed at different levels of clinical trials. This review presents details of CK2 protein, structural insights into adenosine triphosphate binding pocket, current clinical trial candidates and their analogues. Further, it includes the emerging structure-based drug design approaches, chemistry, structure-activity relationship and biological screening of potent and selective CK2 inhibitors. The authors tabulated the details of CK2 co-crystal structures because these co-crystal structures facilitated the structure-guided discovery of CK2 inhibitors. The narrow hinge pocket compared with related kinases provides useful insights into the discovery of CK2 inhibitors.

Autosomal dominant Parkinson's disease caused by the recently identified LRRK2 N1437D mutation in a Chinese family: Clinical features, imaging findings, and functional impact

IntroductionMutations in leucine-rich repeat kinase 2 (LRRK2) are the most common genetic cause of autosomal dominantly inherited Parkinson's disease (PD). Recently, a novel pathogenic variant (N1437D; c.4309A > G; NM_98578) in the LRRK2 gene has been identified in three Chinese families with PD. In this study, we describe a Chinese family with autosomal dominant PD that segregated with the N1437D mutation. A detailed clinical and neuroimaging characterization of the affected family members is reported. We also sought to investigate the functional mechanisms by which the detected mutation could cause PD. MethodsWe characterized the clinical and imaging phenotype of a Chinese pedigree with autosomal dominant PD. We searched for a disease-causing mutation by targeted sequencing and multiple ligation-dependent probe amplification. The functional impact of the mutation was investigated in terms of LRRK2 kinase activity, guanosine triphosphate (GTP) binding, and guanosine triphosphatase (GTPase) activity. ResultsThe disease was found to co-segregate with the LRRK2 N1437D mutation. Patients in the pedigree exhibited typical parkinsonism (age at onset: 54.0 ± 5.9 years). One affected family member – who had evidence of abnormal tau accumulation in the occipital lobe on tau PET imaging – developed PD dementia at follow-up. The mutation markedly increased LRRK2 kinase activity and promoted GTP binding, without affecting GTPase activity. ConclusionsThis study describes the functional impact of a recently identified LRRK2 mutation, N1437D, that causes autosomal dominant PD in the Chinese population. Further research is necessary to investigate the contribution of this mutation to PD in multiple Asian populations.

Multi-omics study of key genes, metabolites, and pathways of periodontitis

ObjectiveThis study aimed to explore the key genes, metabolites, and pathways that influence periodontitis pathogenesis by integrating transcriptomic and metabolomic studies. DesignGingival crevicular fluid samples from periodontitis patients and healthy controls were collected for liquid chromatography/tandem mass-based metabolomics. RNA-seq data for periodontitis and control samples were obtained from the GSE16134 dataset. Differential metabolites and differentially expressed genes (DEGs) between the two groups were then compared. Based on the protein-protein interaction (PPI) network module analysis, key module genes were selected from immune-related DEGs. Correlation and pathway enrichment analyses were performed for differential metabolites and key module genes. A multi-omics integrative analysis was performed using bioinformatic methods to construct a gene-metabolite-pathway network. ResultsFrom the metabolomics study, 146 differential metabolites were identified, which were mainly enriched in the pathways of purine metabolism and Adenosine triphosphate binding cassette transporters (ABC transporters). The GSE16134 dataset revealed 102 immune-related DEGs (458 upregulated and 264 downregulated genes), 33 of which may play core roles in the key modules of the PPI network and are involved in cytokine-related regulatory pathways. Through a multi-omics integrative analysis, a gene-metabolite-pathway network was constructed, including 28 genes (such as platelet derived growth factor D (PDGFD), neurturin (NRTN), and interleukin 2 receptor, gamma (IL2RG)); 47 metabolites (such as deoxyinosine); and 8 pathways (such as ABC transporters). ConclusionPDGFD, NRTN, and IL2RG may be potential biomarkers of periodontitis and may affect disease progression by regulating deoxyinosine to participate in the ABC transporter pathway.

Chronic Rhinosinusitis with Nasal Polyposis in an Adult with Homozygous Variants of Uncertain Significance in ABCA3 Gene

IntroductionThe role of surfactant proteins in pulmonary diseases has been studied extensively. However, little is known about its contribution in diseases involving the nasal mucosa. Chronic rhinosinusitis with nasal polyps (CRSwNP) is a common disease characterized by persistent inflammation of the nasal passage and paranasal sinuses. We present a case of CRSwNP and interstitial lung disease (ILD) with variants of uncertain significance (VUS) in adenosine triphosphate binding cassette subfamily A member 3 (ABCA3) identified as potential genetic etiology. Case DescriptionPatient was a 42-year-old female with chronic obstructive pulmonary disease (COPD) with oxygen-dependance, bronchiectasis, history of hospitalization for bronchiolitis obliterans organizing pneumonia (BOOP) in infancy, who presented for evaluation of CRSwNP and anosmia. Nasal polyps were first discovered around three years of age, requiring more than 25 nasal polypectomies in her lifetime. Evaluation for cystic fibrosis during childhood was unremarkable. She reported only one episode of wheezing and denied aspirin sensitivity. Despite her significant history of BOOP in infancy, the patient remained without pulmonary infections until four years prior to presentation. Since then, she had two episodes of pneumonia and was diagnosed with COPD, leading to chronic oxygen use. Recent spirometry showed severe airflow obstruction, hyperinflation and decreased diffusing capacity for carbon monoxide. Chest CT revealed diffuse bronchiectasis, mucous plugging, and mild ground glass opacity. Patient had negative sweat test and normal alpha-1 antitrypsin level. Genetic testing with Invitae PCD panel and Neonatal Respiratory Distress panel revealed homozygous VUS [c.155C>T (p. Pro52Leu)] in ABCA3 gene. ConclusionOur patient had an atypical course of ILD, remaining asymptomatic most of her life, and CRSwNP. ABCA3 variants are associated with autosomal recessive pulmonary diseases, such as pulmonary surfactant metabolism dysfunction, pediatric ILD and pulmonary fibrosis1. Surfactant phospholipids are known to lower the surface tension in the alveoli, but they also decrease mucus viscosity and help to eliminate inhaled pathogens2,3,4. The pathophysiology of CRSwNP is unclear and the clinical phenotype of the disease varies widely. While the pathogenicity of the variant is not defined, this case raises a possibility that patient’s ABCA3 genotype may contribute to CRSwNP with recalcitrant nasal polyps.

Chemically Fueled Supramolecular Materials.

In biology, the function of many molecules is regulated through nonequilibrium chemical reaction cycles. The prototypical example is the phosphorylation of an amino acid in an enzyme which induces a functional change, e.g., it folds or unfolds, assembles or disassembles, or binds a substrate. Such phosphorylation does not occur spontaneously but requires a phosphorylating agent with high chemical potential (for example, adenosine triphosphate (ATP)) to be converted into a molecule with lower chemical potential (adenosine diphosphate (ADP)). When this energy is used to regulate an assembly, we speak of chemically fueled assemblies; i.e., the molecule with high potential, the fuel, is used to regulate a self-assembly process. For example, the binding of guanosine triphosphate (GTP) to tubulin induces self-assembly. The bound GTP is hydrolyzed to guanosine diphosphate (GDP) upon assembly, which induces tubulin disassembly. The result is a dynamic assembly endowed with unique characteristics, such as time-dependent behavior and the ability to self-heal. These intriguing, unique properties have inspired supramolecular chemists to create similar chemically fueled molecular assemblies from the bottom up. While examples have been designed, they remain scarce partly because chemically fueled reaction cycles are rare and often complex. Thus, we recently developed a carbodiimide-driven reaction cycle that is versatile and easy to use, quantitatively understood, and does not suffer from side reactions. In the reaction cycle, a carboxylate precursor reacts with a carbodiimide to form an activated species like an anhydride or ester. The activated state reacts with water and thereby reverts to its precursor state; i.e., the activated state is deactivated. Effectively, the precursor catalyzes carbodiimides' conversion into waste and forms a transient activated state. We designed building blocks to regulate a range of assemblies and supramolecular materials at the expense of carbodiimide fuel. The simplicity and versatility of the reaction cycles have democratized and popularized the field of chemically fueled assemblies. In this Account, we describe what we have "learned" on our way. We introduce the field exemplified by biological nonequilibrium self-assembly. We describe the design of the carbodiimide-driven reaction cycle. Using examples from our group and others, we offer design rules for the building block's structure and strategies to create the desired morphology or supramolecular materials. The discussed morphologies include fibers, colloids, crystals, and oil- and coacervate-based droplets. We then demonstrate how these assemblies form supramolecular materials with unique material properties like the ability to self-heal. Besides, we discuss the concept of reciprocal coupling in which the assembly exerts feedback on its reaction cycle and we also offer examples of such feedback mechanisms. Finally, we close the Account with a discussion and an outlook on this field. This Account aims to provide our fundamental understanding and facilitate further progress toward conceptually new supramolecular materials.

Gαs slow conformational transition upon GTP binding and a novel Gαs regulator

G proteins are major signaling partners for G protein-coupled receptors (GPCRs). Although stepwise structural changes during GPCR-G protein complex formation and guanosine diphosphate (GDP) release have been reported, no information is available with regard to guanosine triphosphate (GTP) binding. Here, we used a novel Bayesian integrative modeling framework that combines data from hydrogen-deuterium exchange mass spectrometry, tryptophan-induced fluorescence quenching, and metadynamics simulations to derive a kinetic model and atomic-level characterization of stepwise conformational changes incurred by the β2-adrenergic receptor (β2AR)-Gs complex after GDP release and GTP binding. Our data suggest rapid GTP binding and GTP-induced dissociation of Gαs from β2AR and Gβγ, as opposed to a slow closing of the Gαs α-helical domain (AHD). Yeast-two-hybrid screening using Gαs AHD as bait identified melanoma-associated antigen D2 (MAGE D2) as a novel AHD-binding protein, which was also shown to accelerate the GTP-induced closing of the Gαs AHD.

An NMR portrait of functional and dysfunctional allosteric cooperativity in cAMP-dependent protein kinase A.

The cAMP-dependent protein kinase A (PKA) is the archetypical eukaryotic kinase. The catalytic subunit (PKA-C) structure is highly conserved among the AGC-kinase family. PKA-C is a bilobal enzyme with a dynamic N-lobe, harbouring the Adenosine-5'-triphosphate (ATP) binding site and a more rigid helical C-lobe. The substrate-binding groove resides at the interface of the two lobes. A distinct feature of PKA-C is the positive binding cooperativity between nucleotide and substrate. Several PKA-C mutations lead to the development of adenocarcinomas, myxomas, and other rare forms of liver tumours. Nuclear magnetic resonance (NMR) spectroscopy shows that these mutations disrupt the allosteric communication between the two lobes, causing a drastic decrease in binding cooperativity. The loss of cooperativity correlates with changes in substrate fidelity and reduced kinase affinity for the endogenous protein kinase inhibitor (PKI). The similarity between PKI and the inhibitory sequence of the kinase regulatory subunits suggests that the overall mechanism of regulation of the kinase may be disrupted. We surmise that a reduced or obliterated cooperativity may constitute a common trait for both orthosteric and allosteric mutations of PKA-C that may lead to dysregulation and disease.

Signal recognition particle receptor-β (SR-β) coordinates cotranslational N-glycosylation

Proteins destined for the secretory compartment of the cell are cotranslationally translocated into the endoplasmic reticulum. The majority of these proteins are N-glycosylated, a co- and posttranslational modification that ensures proper protein folding, stability, solubility, and cellular localization. Here, we show that the [Formula: see text] subunit of the signal recognition particle receptor (SR) is required for assembly of the N-glycosylation-competent translocon. We report that guanine analog chemical probes identified by high-throughput screening or mutation of the SR-[Formula: see text] guanosine triphosphate binding site cause an N-glycosylation-deficient phenotype. Neither method alters the association of SR-[Formula: see text] with SR-[Formula: see text], but both approaches reduce the association of SR-[Formula: see text] with the oligosaccharyltransferase complex. These experiments demonstrate that SR-[Formula: see text] has a previously unrecognized function coordinating endoplasmic reticulum translation with N-glycosylation.

Morphophysiological, proteomic and metabolomic analyses reveal cadmium tolerance mechanism in common wheat (Triticum aestivum L.).

Soil cadmium (Cd) contamination can reduce wheat yield and quality, thus threatening food security and human health. Herein, morphological physiology, Cd accumulation and distribution, proteomic and metabolomic analyses were performed (using wheat cultivars 'Luomai23' (LM, Cd-sensitive) and 'Zhongyu10' (ZY, Cd-tolerant) at the seedling stage with sand culture) to reveal Cd tolerance mechanism. Cd inhibited wheat growth, caused oxidative stress, hindered carbon and nitrogen metabolism, and altered the quantity and composition of root exudates. The root Cd concentration was lower in ZY than in LM by about 35% under 15μMCd treatments. ZY reduced Cd uptake through root exudation of amino acids and alkaloids. ZY also reduced Cd accumulation through specific up-regulation (twice) of major facilitator superfamily (MFS) proteins. Furthermore, ZY enhanced Cd cell wall fixation and vacuolar compartmentalization by increasing pectin contents, hemicellulose1 contents, and adenosine triphosphate binding cassette subfamily C member 1 (ABCC1) transporter expression, thus reducing the Cd organelle fraction of ZY by about 12% and 44% in root and shoot, respectively, compared with LM. Additionally, ZY had enhanced resilience to Cd due to increased antioxidant capacity, plasma membrane stability, nitrogen metabolism, and endoplasmic reticulum homeostasis, indicating that the increased Cd tolerance could be because of multi-level coordination. These findings provide a reference for exploring the molecular mechanism of Cd tolerance and accumulation, providing a basis for safe utilization of Cd-contaminated soil by breeding Cd-tolerant and low Cd-accumulating wheat varieties.

Preclinical characterization of pirtobrutinib, a highly selective, noncovalent (reversible) BTK inhibitor

AbstractBruton tyrosine kinase (BTK), a nonreceptor tyrosine kinase, is a major therapeutic target for B-cell–driven malignancies. However, approved covalent BTK inhibitors (cBTKis) are associated with treatment limitations because of off-target side effects, suboptimal oral pharmacology, and development of resistance mutations (eg, C481) that prevent inhibitor binding. Here, we describe the preclinical profile of pirtobrutinib, a potent, highly selective, noncovalent (reversible) BTK inhibitor. Pirtobrutinib binds BTK with an extensive network of interactions to BTK and water molecules in the adenosine triphosphate binding region and shows no direct interaction with C481. Consequently, pirtobrutinib inhibits both BTK and BTK C481 substitution mutants in enzymatic and cell-based assays with similar potencies. In differential scanning fluorimetry studies, BTK bound to pirtobrutinib exhibited a higher melting temperature than cBTKi-bound BTK. Pirtobrutinib, but not cBTKis, prevented Y551 phosphorylation in the activation loop. These data suggest that pirtobrutinib uniquely stabilizes BTK in a closed, inactive conformation. Pirtobrutinib inhibits BTK signaling and cell proliferation in multiple B-cell lymphoma cell lines, and significantly inhibits tumor growth in human lymphoma xenografts in vivo. Enzymatic profiling showed that pirtobrutinib was highly selective for BTK in >98% of the human kinome, and in follow-up cellular studies pirtobrutinib retained >100-fold selectivity over other tested kinases. Collectively, these findings suggest that pirtobrutinib represents a novel BTK inhibitor with improved selectivity and unique pharmacologic, biophysical, and structural attributes with the potential to treat B-cell–driven cancers with improved precision and tolerability. Pirtobrutinib is being tested in phase 3 clinical studies for a variety of B-cell malignancies.

Tyk2 Targeting in Immune-Mediated Inflammatory Diseases.

The Janus kinase (Jak)/signal transducer and activating protein (STAT) pathways mediate the intracellular signaling of cytokines in a wide spectrum of cellular processes. They participate in physiologic and inflammatory cascades and have become a major focus of research, yielding novel therapies for immune-mediated inflammatory diseases (IMID). Genetic linkage has related dysfunction of Tyrosine kinase 2 (Tyk2)-the first member of the Jak family that was described-to protection from psoriasis. Furthermore, Tyk2 dysfunction has been related to IMID prevention, without increasing the risk of serious infections; thus, Tyk2 inhibition has been established as a promising therapeutic target, with multiple Tyk2 inhibitors under development. Most of them are orthosteric inhibitors, impeding adenosine triphosphate (ATP) binding to the JH1 catalytic domain-which is highly conserved across tyrosine kinases-and are not completely selective. Deucravacitinib is an allosteric inhibitor that binds to the pseudokinase JH2 (regulatory) domain of Tyk2; this unique mechanism determines greater selectivity and a reduced risk of adverse events. In September 2022, deucravacitinib became the first Tyk2 inhibitor approved for the treatment of moderate-to-severe psoriasis. A bright future can be expected for Tyk2 inhibitors, with newer drugs and more indications to come.

Primer terminal ribonucleotide alters the active site dynamics of DNA polymerase η and reduces DNA synthesis fidelity

DNA polymerases catalyze DNA synthesis with high efficiency, which is essential for all life. Extensive kinetic and structural efforts have been executed in exploring mechanisms of DNA polymerases, surrounding their kinetic pathway, catalytic mechanisms, and factors that dictate polymerase fidelity. Recent time-resolved crystallography studies on DNA polymerase η (Pol η) and β have revealed essential transient events during the DNA synthesis reaction, such as mechanisms of primer deprotonation, separated roles of the three metal ions, and conformational changes that disfavor incorporation of the incorrect substrate. DNA-embedded ribonucleotides (rNs) are the most common lesion on DNA and a major threat to genome integrity. While kinetics of rN incorporation has been explored and structural studies have revealed that DNA polymerases have a steric gate that destabilizes ribonucleotide triphosphate binding, the mechanism of extension upon rN addition remains poorly characterized. Using steady-state kinetics, static and time-resolved X-ray crystallography with Pol η as a model system, we showed that the extra hydroxyl group on the primer terminus does alter the dynamics of the polymerase active site as well as the catalysis and fidelity of DNA synthesis. During rN extension, Pol η error incorporation efficiency increases significantly across different sequence contexts. Finally, our systematic structural studies suggest that the rN at the primer end improves primer alignment and reduces barriers in C2'-endo to C3'-endo sugar conformational change. Overall, our work provides further mechanistic insights into the effects of rN incorporation on DNA synthesis.

Effect of adenosine triphosphate on ribociclib-induced skin toxicity in rats

Purpose Ribociclib is a CDK4/6 inhibitor approved for the treatment of breast cancer; it inhibits the activity of CDK4/6 by competitively binding to adenosine 5'-triphosphate (ATP) binding sites. Although generally well-tolerated, ribociclib has been connected to a number of serious dermatologic complications. This study explored the effects of ATP on ribociclib-induced skin damage. Materials and methods Using a rat model, ATP 25 mg/kg was injected intraperitoneally in the ATP + Ribociclib (ATR) group (n = 6). Distilled water as solvent was applied to the healthy control (HC) group (n = 6) and ribociclib (RCB) group (n = 6). One hour after ATP and solvent administration, ribociclib (200 mg/kg) suspension prepared in distilled water was administered to the stomach by gavage (ATR and RCB groups). This was repeated once a day for 15 d. After that period, biochemical markers were studied in the skin tissues and histopathological evaluations were conducted. Results In the histopathological evaluation of the RCB group, dermal necrosis, degeneration in hair follicles, and pycnosis in keratinocytes were observed. Only mild degeneration was observed in the ATR group; the HC group had a normal histological appearance. The malondialdehyde (MDA) values were significantly higher and the superoxide dismutase (SOD), catalase (CAT), and total glutathione (tGSH) levels were significantly lower in the RCB group in comparison to the HC group (p < .001). ATP reduced the ribociclib-induced increases in the MDA values and decreased the SOD, CAT, and tGSH levels in the ATR group (p < .001). Conclusion ATP may be useful in the treatment of ribociclib-induced skin damage.

Rational design, synthesis and structural characterization of peptides and peptidomimetics to target Hsp90/Cdc37 interaction for treating hepatocellular carcinoma

Heat shock protein 90 (Hsp90) and cell division cycle 37 (Cdc37) work together as a molecular chaperone complex to regulate the activity of a multitude of client protein kinases. These kinases belong to a wide array of intracellular signaling networks that mediate multiple cellular processes including proliferation. As a result, Hsp90 and Cdc37 represent innovative therapeutic targets in various cancers (such as leukemia, multiple myeloma, and hepatocellular carcinoma (HCC)) in which their expression levels are elevated. Conventional small molecule Hsp90 inhibitors act by blocking the conserved adenosine triphosphate (ATP) binding site. However, by targeting less conserved sites in a more specific manner, peptides and peptidomimetics (modified peptides) hold potential as more efficacious and less toxic alternatives to the conventional small molecule inhibitors. Using a rational approach, we herein developed bioactive peptides targeting Hsp90/Cdc37 interaction. A six amino acid linear peptide derived from Cdc37, KTGDEK, was designed to target Hsp90. We used in silico computational docking to first define its mode of interaction, and binding orientation, and then conjugated the peptide with a cell penetrating peptide, TAT, and a fluorescent dye to confirm its ability to colocalize with Hsp90 in HCC cells. Based on the parent linear sequence, we developed a peptidomimetics library of pre-cyclic and cyclic derivatives. These peptidomimetics were evaluated for their binding affinity to Hsp90, and bioactivity in HCC cell lines. Among them, a pre-cyclic peptidomimetic demonstrates high binding affinity and bioactivity in HCC cells, causing reduced cell proliferation that is associated with induction of cell apoptosis, and down-regulation of phosphorylated MEK1/2. Overall, this generalized approach of rational design, structural optimization, and cellular validation of ‘drug-like’ peptidomimetics against Hsp90/Cdc37 offers a feasible and promising way to design novel therapeutic agents for malignancies and other diseases that are dependent on this molecular chaperone complex.