Cavitands as Reaction Vessels and Blocking Groups for Selective Reactions in Water.

The majority of reactions currently performed in the chemical industry take place in organic solvents, compounds that are generally derived from petrochemicals. To promote chemical processes in water, we examined the use of synthetic, deep water-soluble cavitands in the Staudinger reduction of long-chain aliphatic diazides (C8 , C10 , and C12 ). The diazide substrates are taken up by the cavitand in D2 O in folded, dynamic conformations.

The Dependence of Carbohydrate-Aromatic Interaction Strengths on the Structure of the Carbohydrate.

Interactions between proteins and carbohydrates are ubiquitous in biology. Therefore, understanding the factors that determine their affinity and selectivity are correspondingly important. Herein, we have determined the relative strengths of intramolecular interactions between a series of monosaccharides and an aromatic ring close to the glycosylation site in an N-glycoprotein host.

Ring current shifts in (19)F-NMR of membrane proteins.

Fluorine-19 NMR markers are attractive reporter groups for use in studies of complex biomacromolecular systems, in particular also for studies of function-related conformational equilibria and rate processes in membrane proteins. Advantages of (19)F-NMR probes include high sensitivity of the (19)F chemical shifts to variations in the non-covalent environment. Nonetheless, in studies of G protein-coupled receptors (GPCR) we encountered situations where (19)F chemical shifts were not responsive to conformational changes that had been implicated by other methods.

Synthesis and evaluation of duocarmycin SA analogs incorporating the methyl 1,2,8,8a-tetrahydrocyclopropa[c]imidazolo[4,5-e]indol-4-one-6-carboxylate (CImI) alkylation subunit.

The design, synthesis, and evaluation of methyl 1,2,8,8a-tetrahydrocyclopropa[c]imidazolo[4,5-e]indol-4-one-6-carboxylate (CImI) derivatives are detailed representing analogs of duocarmycin SA and yatakemycin containing an imidazole replacement for the fused pyrrole found in the DNA alkylation subunit.

Small-Molecule Fusion Inhibitors Bind the pH-Sensing Stable Signal Peptide-GP2 Subunit Interface of the Lassa Virus Envelope Glycoprotein.

Unlabelled: Arenavirus species are responsible for severe life-threatening hemorrhagic fevers in western Africa and South America. Without effective antiviral therapies or vaccines, these viruses pose serious public health and biodefense concerns. Chemically distinct small-molecule inhibitors of arenavirus entry have recently been identified and shown to act on the arenavirus envelope glycoprotein (GPC) to prevent membrane fusion.

Arylfluorosulfates Inactivate Intracellular Lipid Binding Protein(s) through Chemoselective SuFEx Reaction with a Binding Site Tyr Residue.

Arylfluorosulfates have appeared only rarely in the literature and have not been explored as probes for covalent conjugation to proteins, possibly because they were assumed to possess high reactivity, as with other sulfur(VI) halides. However, we find that arylfluorosulfates become reactive only under certain circumstances, e.g.

Neuronal and Astrocytic Monoacylglycerol Lipase Limit the Spread of Endocannabinoid Signaling in the Cerebellum.

Endocannabinoids are diffusible lipophilic molecules that may spread to neighboring synapses. Monoacylglycerol lipase (MAGL) is the principal enzyme that degrades the endocannabinoid 2-arachidonoylglycerol (2-AG). Using knock-out mice in which MAGL is deleted globally or selectively in neurons and astrocytes, we investigated the extent to which neuronal and astrocytic MAGL limit the spread of 2-AG-mediated retrograde synaptic depression in cerebellar slices.

Current and future treatment of amyloid diseases.

There are more than 30 human proteins whose aggregation appears to cause degenerative maladies referred to as amyloid diseases or amyloidoses. These disorders are named after the characteristic cross-β-sheet amyloid fibrils that accumulate systemically or are localized to specific organs. In most cases, current treatment is limited to symptomatic approaches and thus disease-modifying therapies are needed.

Enhancing protein stability with extended disulfide bonds.

Disulfide bonds play an important role in protein folding and stability. However, the cross-linking of sites within proteins by cysteine disulfides has significant distance and dihedral angle constraints. Here we report the genetic encoding of noncanonical amino acids containing long side-chain thiols that are readily incorporated into both bacterial and mammalian proteins in good yields and with excellent fidelity.

Most neutralizing human monoclonal antibodies target novel epitopes requiring both Lassa virus glycoprotein subunits.

Lassa fever is a severe multisystem disease that often has haemorrhagic manifestations. The epitopes of the Lassa virus (LASV) surface glycoproteins recognized by naturally infected human hosts have not been identified or characterized. Here we have cloned 113 human monoclonal antibodies (mAbs) specific for LASV glycoproteins from memory B cells of Lassa fever survivors from West Africa.

Design of Switchable Chimeric Antigen Receptor T Cells Targeting Breast Cancer.

Chimeric antigen receptor T (CAR-T) cells have demonstrated promising results against hematological malignancies, but have encountered significant challenges in translation to solid tumors. To overcome these hurdles, we have developed a switchable CAR-T cell platform in which the activity of the engineered cell is controlled by dosage of an antibody-based switch. Herein, we apply this approach to Her2-expressing breast cancers by engineering switch molecules through site-specific incorporation of FITC or grafting of a peptide neo-epitope (PNE) into the anti-Her2 antibody trastuzumab (clone 4D5).

Stabilizing the CH2 Domain of an Antibody by Engineering in an Enhanced Aromatic Sequon.

Monoclonal antibodies (mAbs) exhibiting highly selective binding to a protein target constitute a large and growing proportion of the therapeutics market. Aggregation of mAbs results in the loss of their therapeutic efficacy and can result in deleterious immune responses. The CH2 domain comprising part of the Fc portion of Immunoglobulin G (IgG) is typically the least stable domain in IgG-type antibodies and therefore influences their aggregation propensity.

Methamphetamine Vaccines: Improvement through Hapten Design.

Methamphetamine (MA) addiction is a serious public health problem, and current methods to abate addiction and relapse are currently ineffective for mitigating this growing global epidemic. Development of a vaccine targeting MA would provide a complementary strategy to existing behavioral therapies, but this has proven challenging. Herein, we describe optimization of both hapten design and formulation, identifying a vaccine that elicited a robust anti-MA immune response in mice, decreasing methamphetamine-induced locomotor activity.

Engineering a long-acting, potent GLP-1 analog for microstructure-based transdermal delivery.

Antidiabetic treatments aiming to reduce body weight are currently gaining increased interest. Exendin-4, a glucagon-like peptide-1 (GLP-1) receptor agonist administered twice daily via s.c.

AIG1 and ADTRP are atypical integral membrane hydrolases that degrade bioactive FAHFAs.

Enzyme classes may contain outlier members that share mechanistic, but not sequence or structural, relatedness with more common representatives. The functional annotation of such exceptional proteins can be challenging. Here, we use activity-based profiling to discover that the poorly characterized multipass transmembrane proteins AIG1 and ADTRP are atypical hydrolytic enzymes that depend on conserved threonine and histidine residues for catalysis.

Single Hydrogen Bond Donation from Flavin N5 to Proximal Asparagine Ensures FAD Reduction in DNA Photolyase.

The spread of the absorbance of the stable FADH(•) radical (300-700 nm) allows CPD photolyase to highly efficiently form FADH(-), making it functional for DNA repair. In this study, FTIR spectroscopy detected a strong hydrogen bond, from FAD N5-H to the carbonyl group of the Asn378 side chain, that is modulated by the redox state of FAD. The observed characteristic frequency shifts were reproduced in quantum-mechanical models of the flavin binding site, which were then employed to elucidate redox tuning governed by Asn378.

Chemoselective Synthesis of Polysubstituted Pyridines from Heteroaryl Fluorosulfates.

A selection of heteroaryl fluorosulfates were readily synthesized using commercial SO2F2 gas. These substrates are highly efficient coupling partners in the Suzuki reaction. Through judicious selection of Pd catalysts the fluorosulfate functionality is differentiated from bromide and chloride; the order of reactivity being: -Br> -OSO2 F> -Cl.

Conformational dynamics of a membrane protein chaperone enables spatially regulated substrate capture and release.

Membrane protein biogenesis poses enormous challenges to cellular protein homeostasis and requires effective molecular chaperones. Compared with chaperones that promote soluble protein folding, membrane protein chaperones require tight spatiotemporal coordination of their substrate binding and release cycles. Here we define the chaperone cycle for cpSRP43, which protects the largest family of membrane proteins, the light harvesting chlorophyll a/b-binding proteins (LHCPs), during their delivery.

Reactivation of Latent HIV-1 Expression by Engineered TALE Transcription Factors.

The presence of replication-competent HIV-1 -which resides mainly in resting CD4+ T cells--is a major hurdle to its eradication. While pharmacological approaches have been useful for inducing the expression of this latent population of virus, they have been unable to purge HIV-1 from all its reservoirs. Additionally, many of these strategies have been associated with adverse effects, underscoring the need for alternative approaches capable of reactivating viral expression.

Diacylglycerol lipase β inhibition reverses nociceptive behaviour in mouse models of inflammatory and neuropathic pain.

Background And Purpose: Inhibition of diacylglycerol lipase (DGL)β prevents LPS-induced pro-inflammatory responses in mouse peritoneal macrophages. Thus, the present study tested whether DGLβ inhibition reverses allodynic responses of mice in the LPS model of inflammatory pain, as well as in neuropathic pain models.

Experimental Approach: Initial experiments examined the cellular expression of DGLβ and inflammatory mediators within the LPS-injected paw pad.

Crystal Structure of the Oligomeric Form of Lassa Virus Matrix Protein Z.

Unlabelled: The arenavirus matrix protein Z is highly multifunctional and occurs in both monomeric and oligomeric forms. The crystal structure of a dodecamer of Z from Lassa virus, presented here, illustrates a ring-like structure with a highly basic center. Mutagenesis demonstrates that the dimeric interface within the dodecamer and a Lys-Trp-Lys triad at the center of the ring are important for oligomerization.

Host-Primed Ebola Virus GP Exposes a Hydrophobic NPC1 Receptor-Binding Pocket, Revealing a Target for Broadly Neutralizing Antibodies.

Unlabelled: The filovirus surface glycoprotein (GP) mediates viral entry into host cells. Following viral internalization into endosomes, GP is cleaved by host cysteine proteases to expose a receptor-binding site (RBS) that is otherwise hidden from immune surveillance. Here, we present the crystal structure of proteolytically cleaved Ebola virus GP to a resolution of 3.

Glycation Reactivity of a Quorum-Sensing Signaling Molecule.

Reported herein is that (4S)-4,5-dihydroxy-2,3-pentanedione (DPD) can undergo a previously undocumented non-enzymatic glycation reaction. Incubation of DPD with viral DNA or the antibiotic gramicidin S resulted in significant biochemical alterations. A protein-labeling method was consequently developed that facilitated the identification of unrecognized glycation targets of DPD in a prokaryotic system.

High-Resolution Mapping of the Folding Transition State of a WW Domain.

Fast-folding WW domains are among the best-characterized systems for comparing experiments and simulations of protein folding. Recent microsecond-resolution experiments and long duration (totaling milliseconds) single-trajectory modeling have shown that even mechanistic changes in folding kinetics due to mutation can now be analyzed. Thus, a comprehensive set of experimental data would be helpful to benchmark the predictions made by simulations.

Combatting Synthetic Designer Opioids: A Conjugate Vaccine Ablates Lethal Doses of Fentanyl Class Drugs.

Fentanyl is an addictive prescription opioid that is over 80 times more potent than morphine. The synthetic nature of fentanyl has enabled the creation of dangerous "designer drug" analogues that escape toxicology screening, yet display comparable potency to the parent drug. Alarmingly, a large number of fatalities have been linked to overdose of fentanyl derivatives.