Expression, purification, and biophysical analysis of a part of the C-terminal domain of human hypoxia inducible factor-2α (HIF-2α).

Hypoxia inducible factor 2α (HIF-2α) is a member of the basic helix-loop-helix(bHLH)-Per-Arnt-Sim (PAS) family of transcription factors. It is overexpressed in several cancers, associated with poor prognosis of the patients and resistance to treatment. Here, we study the residues 366-704 of the C-terminal end of human HIF-2α, which contains the N-transcriptional activation domain (NTAD), the oxygen-dependent degradation domain (ODD), and a part of the inhibitory domain (IH).

Identification of Small-Molecule Antagonists Targeting the Growth Hormone Releasing Hormone Receptor (GHRHR).

The growth hormone-releasing hormone receptor (GHRHR) belongs to Class B1 of G protein-coupled receptors (GPCRs). Class B1 GPCR peptides such, as growth hormone-releasing hormone (GHRH), have been proposed to bind in a two-step model, where first the C-terminal region of the peptide interacts with the extracellular domain of the receptor and, subsequently, the N-terminus interacts with the seven transmembrane domain of the receptor, resulting in activation. The GHRHR has recently been highlighted as a promising drug target toward several types of cancer and has been shown to be overexpressed in prostate, breast, pancreatic, and ovarian cancer.

Identification of Novel Biomarkers in Late Preterm Neonates with Respiratory Distress Syndrome (RDS) Using Urinary Metabolomic Analysis.

Urine metabolomics is gaining traction as a means of identifying metabolic signatures associated with health and disease states. Thirty-one (31) late preterm (LP) neonates admitted to the neonatal intensive care unit (NICU) and 23 age-matched healthy LPs admitted to the maternity ward of a tertiary hospital were included in the study. Proton nuclear magnetic resonance (H NMR) spectroscopy was employed for urine metabolomic analysis on the 1st and 3rd days of life of the neonates.

Structural and Biochemical Characterization of the Human Angiogenin-Proliferating Cell Nuclear Antigen Interaction.

The molecular details of the interaction between human angiogenin (hAng) and proliferating cell nuclear antigen (PCNA) have been investigated by isothermal titration calorimetry (ITC), mutagenesis, and NMR spectroscopy. The two proteins were shown to interact directly through immunoprecipitation studies of hAng with PCNA , and their interaction was quantified by ITC, obtaining information on stoichiometry, enthalpy, entropy, and binding kinetics of the association. The hAng-PCNA association is strong, with a value of 126 nM.

E2 Partner Tunes the Ubiquitylation Specificity of Arkadia E3 Ubiquitin Ligase.

Arkadia () is a positive regulator of the TGF- signaling that mediates the proteasome-dependent degradation of negative factors of the pathway. It is classified as an E3 ubiquitin ligase and a SUMO-targeted ubiquitin ligase (STUBL), implicated in various pathological conditions including cancer and fibrosis. The enzymatic (ligase) activity of Arkadia is located at its C-terminus and involves the RING domain.

NMR Analysis Suggests Synergy between the RRM2 and the Carboxy-Terminal Segment of Human La Protein in the Recognition and Interaction with HCV IRES.

The La protein (lupus antigen) is a ubiquitous RNA-binding protein found in all human cells. It is mainly localized in the nucleus, associates with all RNA polymerase III (Pol III) transcripts, as the first factor they interact with, and modulates subsequent processing events. Export of La to the cytoplasm has been reported to stimulate the decoding of specific cellular and viral mRNAs through IRES-dependent (Internal ribosome entry site) binding and translation.

Peptide inhibitors of angiotensin-I converting enzyme based on angiotensin (1-7) with selectivity for the C-terminal domain.

The renin-angiotensin system (RAS) is a key regulator of human arterial pressure. Several of its effects are modulated by angiotensin II, an octapeptide originating from the action of angiotensin-I converting enzyme (ACE) on the decapeptide angiotensin-I. ACE possess two active sites (nACE and cACE) that have their own kinetic and substrate specificities.

H, C, N backbone resonance assignment of apo and ADP-ribose bound forms of the macro domain of Hepatitis E virus through solution NMR spectroscopy.

The genome of Hepatitis E virus (HEV) is 7.2 kilobases long and has three open reading frames. The largest one is ORF1, encoding a non-structural protein involved in the replication process, and whose processing is ill-defined.

NMR-based metabolic profiling of children with premature adrenarche.

Introduction: Premature adrenarche (PA) for long time was considered a benign condition but later has been connected to various diseases in childhood and adulthood which remains controversial.

Objective: To investigate the effect of premature adrenarche on the metabolic phenotype, and correlate the clinical and biochemical data with the metabolic profile of children with PA.

Methods: Nuclear magnetic resonance (NMR)-based untargeted and targeted metabolomic approach in combination with multivariate and univariate statistical analysis applied to study the metabolic profiles of children with PA.

Unveiling the Essential Role of Arkadia's Non-RING Elements in the Ubiquitination Process.

Arkadia is a positive regulator of the TGF-SMAD2/3 pathway, acting through its C-terminal RING-H2 domain and targeting for degradation of its negative regulators. Here we explore the role of regions outside the RING domain (non-RING elements) of Arkadia on the E2-E3 interaction. The contribution of the non-RING elements was addressed using Arkadia RING 68 aa and Arkadia 119 aa polypeptides.

Comprehensive Fragment Screening of the SARS-CoV-2 Proteome Explores Novel Chemical Space for Drug Development.

SARS-CoV-2 (SCoV2) and its variants of concern pose serious challenges to the public health. The variants increased challenges to vaccines, thus necessitating for development of new intervention strategies including anti-virals. Within the international Covid19-NMR consortium, we have identified binders targeting the RNA genome of SCoV2.

NMR study of human macroPARPs domains: H, N and C resonance assignment of hPARP14 macro domain 2 in the free and the ADPr bound state.

hPARP14 is a human ADP-ribosyl-transferase (ART) that belongs to the macroPARPs family, together with hPARP9 and hPARP15. It contains a tandem of three macro domains (MD) while each of them has different properties. The first one, namely MD1, has not been reported to exhibit a high binding affinity for ADP-ribose (ADPr) in contrast to the following two (MD2 and MD3).

Backbone and side chain NMR assignment of the heme-nitric oxide/oxygen binding (H-NOX) domain from Nostoc punctiforme.

Soluble guanylate cyclase (sGC) is considered as the primary NO receptor across several known eukaryotes. The main interest regarding the biological role and its function, focuses on the H-NOX domain of the β1 subunit. This domain in its active form bears a ferrous b type heme as prosthetic group, which facilitates the binding of NO and other diatomic gases.

Binding Adaptation of GS-441524 Diversifies Macro Domains and Downregulates SARS-CoV-2 de-MARylation Capacity.

Viral infection in cells triggers a cascade of molecular defense mechanisms to maintain host-cell homoeostasis. One of these mechanisms is ADP-ribosylation, a fundamental post-translational modification (PTM) characterized by the addition of ADP-ribose (ADPr) on substrates. Poly(ADP-ribose) polymerases (PARPs) are implicated in this process and they perform ADP-ribosylation on host and pathogen proteins.

Mapping of the sGC Stimulator BAY 41-2272 Binding Site on H-NOX Domain and Its Regulation by the Redox State of the Heme.

Soluble guanylate cyclase (sGC) is the main receptor of nitric oxide (NO) and by converting GTP to cGMP regulates numerous biological processes. The 1 subunit of the most abundant, 11 heterodimer, harbors an N-terminal domain called H-NOX, responsible for heme and NO binding and thus sGC activation. Dysfunction of the NO/sGC/cGMP axis is causally associated with pathological states such as heart failure and pulmonary hypertension.

NMR-Based Metabolomics in Differential Diagnosis of Chronic Kidney Disease (CKD) Subtypes.

Chronic Kidney Disease (CKD) is considered as a major public health problem as it can lead to end-stage kidney failure, which requires replacement therapy. A prompt and accurate diagnosis, along with the appropriate treatment, can delay CKD's progression, significantly. Herein, we sought to determine whether CKD etiology can be reflected in urine metabolomics during its early stage.

Losartan Interactions with 2-Hydroxypropyl-β-CD.

Losartan potassium salt (LSR) is a well-known antihypertensive drug with proven beneficial effects on human health. Its formulation with the non-toxic 2-hydroxypropyl-β-cyclodextrin (2-HP-β-CD) could improve its pharmacological profile. Thus, its molecular interactions are studied using a combination of Differential Scanning Calorimetry (DSC), Nuclear Magnetic Resonance (NMR) and Molecular Dynamics (MD).

Impact of a Single Nucleotide Polymorphism on the 3D Protein Structure and Ubiquitination Activity of E3 Ubiquitin Ligase Arkadia.

Single nucleotide polymorphisms (SNPs) are genetic variations which can play a vital role in the study of human health. SNP studies are often used to identify point mutations that are associated with diseases. Arkadia (RNF111) is an E3 ubiquitin ligase that enhances transforming growth factor-beta (TGF-) signaling by targeting negative regulators for degradation.

Metabolite and Bioactive Compounds Profiling of Meteora Sea Buckthorn Berries through High-Resolution NMR Analysis.

Sea buckthorn berries ( L.) (SB) are considered as a fruit with a high nutritional value with a plethora of bioactive ingredients. The present work focusses on the analysis of the whole NMR metabolic profile of SB berries grown in an organic orchard of Meteora/Greece.

Replacement of heme by soluble guanylate cyclase (sGC) activators abolishes heme-nitric oxide/oxygen (H-NOX) domain structural plasticity.

The gasotransmitter nitric oxide (NO) is a critical endogenous regulator of homeostasis, in major part via the generation of cGMP (cyclic guanosine monophosphate) from GTP (guanosine triphosphate) by NO's main physiological receptor, the soluble guanylate cyclase (sGC). sGC is a heterodimer, composed of an α1 and a β1 subunit, of which the latter contains the heme-nitric oxide/oxygen (H-NOX) domain, responsible for NO recognition, binding and signal initiation. The NO/sGC/cGMP axis is dysfunctional in a variety of diseases, including hypertension and heart failure, especially since oxidative stress results in heme oxidation, sGC unresponsiveness to NO and subsequent degradation.

3D Reconstitution of the Neural Stem Cell Niche: Connecting the Dots.

Neural stem cells (NSCs) are important constituents of the nervous system, and they become constrained in two specific regions during adulthood: the subventricular zone (SVZ) and the subgranular zone (SGZ) of the dentate gyrus in the hippocampus. The SVZ niche is a limited-space zone where NSCs are situated and comprised of growth factors and extracellular matrix (ECM) components that shape the microenvironment of the niche. The interaction between ECM components and NSCs regulates the equilibrium between self-renewal and differentiation.

Nontargeted H NMR fingerprinting and multivariate statistical analysis for traceability of Greek PDO Vostizza currants.

In this study, non-targeted H NMR fingerprinting was used in combination with multivariate statistical analyses for the classification of Greek currants based on their geographical origins (Aeghion, Nemea, Kalamata, Zante, and Amaliada). As classification techniques, Principal Component Analysis (PCA) and Partial Least Squares Discriminant Analysis (PLS-DA) were carried out. To elucidate different components according to PDO (Protected Designation of Origin), products from Aeghion (Vostizza) were statistically compared with each one of the four other regions.

Three-Dimensional Cell Metabolomics Deciphers the Anti-Angiogenic Properties of the Radioprotectant Amifostine.

Aberrant angiogenesis is a hallmark for cancer and inflammation, a key notion in drug repurposing efforts. To delineate the anti-angiogenic properties of amifostine in a human adult angiogenesis model via 3D cell metabolomics and upon a stimulant-specific manner, a 3D cellular angiogenesis assay that recapitulates cell physiology and drug action was coupled to untargeted metabolomics by liquid chromatography-mass spectrometry and nuclear magnetic resonance spectroscopy. The early events of angiogenesis upon its most prominent stimulants (vascular endothelial growth factor-A or deferoxamine) were addressed by cell sprouting measurements.