ACS Bio & Med Chem Au最新文献

A myriad of biological processes are facilitated by ligand–receptor interactions. The low affinities of these interactions are typically enhanced by multivalent engagements to promote binding. However, each biological interaction requires a unique display and orientation of ligands. Therefore, the availability and diversity of synthetic multivalent probes are invaluable to the investigation of ligand–receptor binding interactions. Here, we report glycopolymers prepared from bicyclo[4.2.0]oct-6-ene-7-carboxamide and 4,7-dihydro-1,3-dioxepin or cyclohexene. These glycopolymers, synthesized by alternating ring-opening metathesis polymerization, display precise ligand spacing as well as the option of a hydrophobic or acetal-functionalized polymer backbone. Small-angle X-ray scattering (SAXS) data analysis revealed that these [4.2.0] glycopolymers adopted distinct conformations in solution. In aqueous media, [4.2.0]-dioxepin glycopolymers formed swollen polymer chains with rod-like, flexible structures while [4.2.0]-cyclohexene glycopolymers assumed compact, globular structures. To illustrate how these glycopolymers could aid in the exploration of ligand–receptor interactions, we incorporated the [4.2.0] glycopolymers into a biological assay to assess their potential as activators of acrosomal exocytosis (AE) in mouse sperm. The results of the biological assay confirmed that the differing structures of the [4.2.0] glycopolymers would evoke distinct biological responses; [4.2.0]-cyclohexene glycopolymers induced AE in mouse sperm while [4.2.0]-dioxepin glycopolymers did not. Herein, we provide two options for glycopolymers with low to moderate molecular weight dispersities and low cytotoxicity that can be implemented into biological assays based on the desired hydrophobicity, rigidity, and structural conformation of the polymer probe.

Carbohydrate recognition is imperative for the induction of sperm acrosomal exocytosis (AE), an essential phenomenon in mammalian fertilization. In mouse sperm, polynorbornene 100-mers displaying fucose or mannose moieties were effective at inducing AE. In contrast, glycopolymers exhibiting glucose sugars resulted in no AE activation. To further elucidate the role of ligand density on the activation of AE in mouse sperm, a triple-stain flow cytometry assay was employed to determine the efficacy of polynorbornene block copolymers with barbell-like sequences as initiators of AE. Triblock (ABA or ABC) copolymers were synthesized by ring-opening metathesis polymerization (ROMP) with one or two activating sugars, mannose or fucose, and one nonactivating sugar, glucose. The active ligand fractions in the polymers varied from 10, 20, or 40%. Simultaneously, random copolymers comprising 20% activating ligands were prepared to confirm the importance of ligand positionality in AE activation in mouse sperm. Polynorbornene 100-mers possessing two 10-mer blocks of activating sugars were the most effective copolymers at inducing AE with levels of AE comparable to their homopolymer counterparts and more effective than their random analogues. Small-angle X-ray scattering (SAXS) was then performed to verify that there were no differences in the conformations of the glycopolymers contributing to their varying AE activity. SAXS data analysis confirmed that all of the glycopolymers assumed semiflexible cylindrical structures with similar radii and Kuhn lengths. These findings suggest that the overall ligand density of the sugar moieties in the polymer is less important than the positionality of short blocks of high-density ligands for AE activation in mouse sperm.

Synthetic modification of oligodeoxynucleotides (ODNs) via conjugation to nucleic acid binding small molecules can improve hybridization and pharmacokinetic properties. In the present study, five Hoechst 33258 derived benzimidazoles were conjugated to T rich ODNs and their hybridization effectiveness was tested. Thermal denaturation studies revealed significant stabilization of complementary duplexes by ODN-benzimidazole conjugates, with the extent of stabilization being highly dependent on the length of the linker between DNA and benzimidazole. The increases in thermal stability were determined to be due to the binding of the benzimidazole moiety to the duplex. Circular dichroism and molecular modeling studies provided insights toward the influence of conjugation on duplex structure and how linker length impacts placement of the benzimidazole moiety in the minor groove. Furthermore, thermal denaturation studies with the complementary strand containing a single base mismatch or being RNA revealed that covalent conjugation of benzimidazoles to an ODN also enhances the sequence specificity. The fundamental studies reported herein provide a strategy to improve the stability and specificity properties of the ODN probes, which can be of use for targeting and diagnostics applications.

The biomolecular corona, a complex layer of biological molecules, envelops nanoparticles (NPs) upon exposure to biological fluids including blood. This dynamic interface is pivotal for the advancement of nanomedicine, particularly in areas of therapy and diagnostics. In situ analysis of the biomolecular corona is crucial, as it can substantially improve our ability to accurately predict the biological fate of nanomedicine and, therefore, enable development of more effective, safe, and precisely targeted nanomedicines. Despite its importance, the repertoire of techniques available for in situ analysis of the biomolecular corona is surprisingly limited. This tutorial review provides an overview of the available techniques for in situ analysis of biomolecular corona with a particular focus on exploring both the advantages and the limitations inherent in the use of field-flow fractionation (FFF) for in situ analysis of the biomolecular corona. It delves into how FFF can unravel the complexities of the corona, enhancing our understanding and guiding the design of next-generation nanomedicines for medical use.

NS1 in flaviviruses is the only nonstructural protein that is secretory and interacts with different cellular components of the host cell membrane. NS1 is localized in the ER as a dimer to facilitate viral replication. Crystal structures of NS1 homologues from zika (ZIKV) and dengue (DENV) viruses have revealed the organization of different domains in NS1 dimers. The β-roll and the connector and intertwined loop regions of wing domains of NS1 have been shown to interact with the membranes. In this study, we have performed multiple molecular dynamics (MD) simulations of ZIKV and DENV NS1 systems in apo and in POPE bilayers with different cholesterol concentrations (0, 20 and 40%). The NS1 protein was placed just above the membrane surface, and for each NS1-membrane system two to three independent simulations with 600 ns production run were performed. At the end of the production runs, ZIKV NS1 inserts deeper inside the membrane compared to the DENV counterpart. Unlike ZIKV NS1, the orientation of DENV NS1 is asymmetric in which one of the chains in the dimer interacts with the membrane while the other is more exposed to the solvent. The β-roll region in ZIKV NS1 penetrates beyond the headgroup region and interacts with the lipid acyl chains while the C-terminal region barely interacts with the headgroup. Specific residues in the intertwined region deeply penetrate inside the membrane. The role of charged and aromatic residues of ZIKV NS1 in strongly interacting with the membrane components is revealed. The presence of cholesterol affects the extent of insertion in the membrane and interaction of individual residues. Overall, membrane-binding properties of ZIKV NS1 significantly differ from its counterpart in DENV. The differences found in the binding and insertion of NS1 can be used to design drugs and novel antibodies that can be flavivirus specific.

Neurodegeneration with brain iron accumulation (NBIA) is a group of neurodegenerative diseases that are typically caused by a monogenetic mutation, leading to development of disordered movement symptoms such as dystonia, hyperreflexia, etc. Brain iron accumulation can be diagnosed through MRI imaging and is hypothesized to be the cause of oxidative stress, leading to the degeneration of brain tissue. There are four main types of NBIA: pantothenate kinase-associated neurodegeneration (PKAN), PLA2G6-associated neurodegeneration (PLAN), mitochondrial membrane protein-associated neurodegeneration (MKAN), and beta-propeller protein-associated neurodegeneration (BPAN). There are no causative therapies for these diseases, but iron chelators have been shown to have potential toward treating NBIA. Three chelators are investigated in this Review: deferoxamine (DFO), desferasirox (DFS), and deferiprone (DFP). DFO has been investigated to treat neurodegenerative diseases such as Alzheimer’s disease (AD) and Parkinson’s disease (PD); however, dose-related toxicity in these studies, as well as in PKAN studies, have shown that the drug still requires more development before it can be applied toward NBIA cases. Iron chelation therapies other than the ones currently in clinical use have not yet reached clinical studies, but they may possess characteristics that would allow them to access the brain in ways that current chelators cannot. Intranasal formulations are an attractive dosage form to study for chelation therapy, as this method of delivery can bypass the blood-brain barrier and access the CNS. Gene therapy differs from iron chelation therapy as it is a causal treatment of the disease, whereas iron chelators only target the disease progression of NBIA. Because the pathophysiology of NBIA diseases is still unclear, future courses of action should be focused on causative treatment; however, iron chelation therapy is the current best course of action.

Phytohormones have significant roles in redox metabolism, inflammatory responses, and cellular survival mechanisms within the microenvironment of the mammalian brain. Herein, we identified the mammalian molecular targets of three representative strigolactone (SL) analogues structurally derived from apocarotenoids and the functional equivalent of plant hormones. All tested SL analogues have an inhibitory effect on NLRP3 inflammasome-mediated IL-1β release in murine microglial cells. However, IND and EGO10 became prominent among them due to their high potency at low micromolar doses. All SL analogues dose-dependently suppressed the release and expression of proinflammatory factors. For EGO10 and IND, IC₅₀ values for iNOS-associated NO secretion were as low as 1.72 and 1.02 μM, respectively. In silico analyses revealed that (S)-EGO10 interacted with iNOS, NLRP3, and Keap1 ligands with the highest binding affinities among all stereoisomeric SL analogues. Although all compounds were effective in microglial Mox phenotype polarization, 4-Br-debranone exhibited a differential pattern for upregulating Nrf2-driven downstream enzymes.

The persistence of neurodegenerative diseases has necessitated the development of new strategies to monitor protein homeostasis (proteostasis). Previous efforts in our laboratory have focused on the development of fluorogenic strategies to observe the onset and progression of proteostatic stress. These works utilized solvatochromic and viscosity sensitive fluorophores to sense protein folded states, enabling stressor screening with an increase in the emission intensity upon aggregation. In this work, we present a novel, high-fidelity assay to detect perturbations of cellular proteostasis, where the fluorescence intensity decreases with the onset of proteostatic stress. Utilizing a fluorogenic, hydroxymethyl silicon-rhodamine probe to differentiate between protein folded states, we establish the validity of this technology in living cells by demonstrating a two-fold difference in fluorescence intensity between unstressed and stressed conditions.