N 1,N 2-Bis­(2,6-di­methyl­phen­yl)-N 1-hydroxyformamidine N,N′-bis­(2,6-dimethyl­phen­yl)-N-oxidoformamidinium dichloro­methane solvate

The title compound, 2C17H20N2O·CH2Cl2, was obtained by N-oxidation of the parent formamidine with m-chloro-peroxy­benzoic acid (m-CPBA). This is the first use of the above-mentioned synthetic route for the preparation of hydroxy­amidines. The title compound crystallizes as a cyclic dimer resulting from the presence of O—H⋯O and N—H⋯N hydrogen bonds

Desquamative Gingivitis: Early Presenting Symptom of Mucocutaneous Disease

Desquamation of the gingiva is a sign that may be encountered in clinical practice. Various diseases can affect the gingival tissues. Mild desquamation that is localized may be associated with mechanical irritation or induced by trauma. Moderate to severe generalized desquamation associated with ulceration and erythema may be indicative of a more serious systemic condition. Although often overlooked, mucocutaneous diseases frequently present with gingival desquamation as an early presenting symptom. The most common mucocutaneous diseases that affect the oral cavity are lichen planus, pemphigus, and mucous membrane pemphigoid. This article reviews the etiology, signs and symptoms, and therapies for these disorders. Increased knowledge of mucocutaneous diseases can help the clinician recognize these disorders and enable the patient to receive appropriate therapy

Thio-linked UDP-peptide conjugates as O-GlcNAc transferase inhibitors

O-GlcNAc transferase (OGT) is an essential glycosyltransferase that installs the O-GlcNAc post-translational modification on the nucleocytoplasmic proteome. We report the development of S-linked UDP–peptide conjugates as potent bisubstrate OGT inhibitors. These compounds were assembled in a modular fashion by photoinitiated thiol–ene conjugation of allyl-UDP and optimal acceptor peptides in which the acceptor serine was replaced with cysteine. The conjugate VTPVC­(S-propyl-UDP)­TA (<i>K</i>_i = 1.3 μM) inhibits the OGT activity in HeLa cell lysates. Linear fusions of this conjugate with cell penetrating peptides were explored as prototypes of cell-penetrant OGT inhibitors. A crystal structure of human OGT with the inhibitor revealed mimicry of the interactions seen in the pseudo-Michaelis complex. Furthermore, a fluorophore-tagged derivative of the inhibitor works as a high affinity probe in a fluorescence polarimetry hOGT assay

Gold nanoparticles delivery in mammalian live cells: a critical review

Functional nanomaterials have recently attracted strong interest from the biology community, not only as potential drug delivery vehicles or diagnostic tools, but also as optical nanomaterials. This is illustrated by the explosion of publications in the field with more than 2,000 publications in the last 2 years (4,000 papers since 2000; from ISI Web of Knowledge, ‘nanoparticle and cell’ hit). Such a publication boom in this novel interdisciplinary field has resulted in papers of unequal standard, partly because it is challenging to assemble the required expertise in chemistry, physics, and biology in a single team. As an extreme example, several papers published in physical chemistry journals claim intracellular delivery of nanoparticles, but show pictures of cells that are, to the expert biologist, evidently dead (and therefore permeable). To attain proper cellular applications using nanomaterials, it is critical not only to achieve efficient delivery in healthy cells, but also to control the intracellular availability and the fate of the nanomaterial. This is still an open challenge that will only be met by innovative delivery methods combined with rigorous and quantitative characterization of the uptake and the fate of the nanoparticles. This review mainly focuses on gold nanoparticles and discusses the various approaches to nanoparticle delivery, including surface chemical modifications and several methods used to facilitate cellular uptake and endosomal escape. We will also review the main detection methods and how their optimum use can inform about intracellular localization, efficiency of delivery, and integrity of the surface capping

Report of the 12th Liaison Meeting

The 12th Liaison meeting was held in Brussels on 8th and 9th October 2015 to address the following Terms of Reference: TOR 1. Discussion on possible follow-­‐‑up to the main outputs/recommendations of: • The 2015 RCMs -­‐‑ specific recommendations addressed to the Liaison Meeting • PGECON, PGDATA, PGMed – outcomes and recommendations from their 2015 meeting • STECF EWG and STECF Plenary -­‐‑ outcomes and recommendations from their 2015 meetings • Data end users (ICES, STECF, RFMOs – GFCM, IATTC, ICCAT, IOTC, WCPFC, NAFO, SPRFMO, CECAF, WECAFC) TOR2. End user feedback on data transmission and related issues • Discuss feedback received from data end-­‐‑users on data transmission: main issues and possible harmonization of end user feedback to the Commission • JRC data transmission IT platform: experience gained and future steps • Discuss best practices on automatization of data upload by MS: data validation tools used by end users • Discussion on new set-­‐‑up for STECF evaluation of AR2014 & data transmission 2014 used in 2015 – continue like this next year? • Harmonisation and dissemination of DCF metadata: codelists, metiers, nomenclatures, best practices, standards • RCM data calls – overview of how MS responded TOR 3. Regional cooperation • Call for proposals MARE/2014/19 'ʹStrengthening Regional Cooperation in the area of fisheries data collection– state of play'ʹ. Presentation by a representative of the two RCG grants and discussions by LM thereafter. What should be the way forward? • Regional databases • Overview of use of the Regional Databases for RCMs in 2015 and problems identified • Other developments (RDB trainings in 2015, RDB Med&BS development) • Changes for the future – any recommendations from the LM? • Future role of RCMs and DCF-­‐‑related meetings: best practices, coordination, cohesion and common structure in line with emerging needs of DCF TOR 4. EU MAP • Discuss recommendations/ output of RCMs: List of proposed stocks, landing obligation, metiers • Discuss design-­‐‑based sampling in relation to DCF: does it fulfil DCF requirements? TOR 5. Availability of data • Overview of latest developments (DCF Database Feasibility Study and plans for a follow-­‐‑up study to this) TOR 6. AOB • Agree on a list of recommendations relating to DCF (that MS will need to report on in their AR2015) – COM will provide a compilation of proposed recommendations from LM & STECF Plenaries in 2014 as input • Prepare a list of recommended meetings for 2016 as guidance for MS • Review and prioritize DCF-­‐‑related study proposals from RCMs, PGECON, EGs etc • ICES update on workshop on concurrent sampling and plans to re-­‐‑evaluate survey

Combinatorial targeting and discovery of ligand-receptors in organelles of mammalian cells

Phage display screening allows the study of functional protein–protein interactions at the cell surface, but investigating intracellular organelles remains a challenge. Here we introduce internalizing-phage libraries to identify clones that enter mammalian cells through a receptor-independent mechanism and target-specific organelles as a tool to select ligand peptides and identify their intracellular receptors. We demonstrate that penetratin, an antennapedia-derived peptide, can be displayed on the phage envelope and mediate receptor-independent uptake of internalizing phage into cells. We also show that an internalizing-phage construct displaying an established mitochondria-specific localization signal targets mitochondria, and that an internalizing-phage random peptide library selects for peptide motifs that localize to different intracellular compartments. As a proof-of-concept, we demonstrate that one such peptide, if chemically fused to penetratin, is internalized receptor-independently, localizes to mitochondria, and promotes cell death. This combinatorial platform technology has potential applications in cell biology and drug development

Truncated and Helix-Constrained Peptides with High Affinity and Specificity for the cFos Coiled-Coil of AP-1

Protein-based therapeutics feature large interacting surfaces. Protein folding endows structural stability to localised surface epitopes, imparting high affinity and target specificity upon interactions with binding partners. However, short synthetic peptides with sequences corresponding to such protein epitopes are unstructured in water and promiscuously bind to proteins with low affinity and specificity. Here we combine structural stability and target specificity of proteins, with low cost and rapid synthesis of small molecules, towards meeting the significant challenge of binding coiled coil proteins in transcriptional regulation. By iteratively truncating a Jun-based peptide from 37 to 22 residues, strategically incorporating i-->i+4 helix-inducing constraints, and positioning unnatural amino acids, we have produced short, water-stable, alpha-helical peptides that bind cFos. A three-dimensional NMR-derived structure for one peptide (24) confirmed a highly stable alpha-helix which was resistant to proteolytic degradation in serum. These short structured peptides are entropically pre-organized for binding with high affinity and specificity to cFos, a key component of the oncogenic transcriptional regulator Activator Protein-1 (AP-1). They competitively antagonized the cJun–cFos coiled-coil interaction. Truncating a Jun-based peptide from 37 to 22 residues decreased the binding enthalpy for cJun by ~9 kcal/mol, but this was compensated by increased conformational entropy (TDS ≤ 7.5 kcal/mol). This study demonstrates that rational design of short peptides constrained by alpha-helical cyclic pentapeptide modules is able to retain parental high helicity, as well as high affinity and specificity for cFos. These are important steps towards small antagonists of the cJun-cFos interaction that mediates gene transcription in cancer and inflammatory diseases

Non-Metabolic Membrane Tubulation and Permeability Induced by Bioactive Peptides

BACKGROUND: Basic cell-penetrating peptides are potential vectors for therapeutic molecules and display antimicrobial activity. The peptide-membrane contact is the first step of the sequential processes leading to peptide internalization and cell activity. However, the molecular mechanisms involved in peptide-membrane interaction are not well understood and are frequently controversial. Herein, we compared the membrane activities of six basic peptides with different size, charge density and amphipaticity: Two cell-penetrating peptides (penetratin and R9), three amphipathic peptides and the neuromodulator substance P. METHODOLOGY/PRINCIPAL FINDINGS: Experiments of X ray diffraction, video-microscopy of giant vesicles, fluorescence spectroscopy, turbidimetry and calcein leakage from large vesicles are reported. Permeability and toxicity experiments were performed on cultured cells. The peptides showed differences in bilayer thickness perturbations, vesicles aggregation and local bending properties which form lipidic tubular structures. These structures invade the vesicle lumen in the absence of exogenous energy. CONCLUSIONS/SIGNIFICANCE: We showed that the degree of membrane permeabilization with amphipathic peptides is dependent on both peptide size and hydrophobic nature of the residues. We propose a model for peptide-induced membrane perturbations that explains the differences in peptide membrane activities and suggests the existence of a facilitated “physical endocytosis,” which represents a new pathway for peptide cellular internalization

A high-throughput synthetic platform enables the discovery of proteomimetic cell penetrating peptides and bioportides

Collectively, cell penetrating peptide (CPP) vectors and intrinsically active bioportides possess tremendous potential for drug delivery applications and the discrete modulation of intracellular targets including the sites of protein–protein interactions (PPIs). Such sequences are usually relatively short (< 25 AA), polycationic in nature and able to access the various intracellular compartments of eukaryotic cells without detrimental influences upon cellular biology. The high-throughput platform for bioportide discovery described herein exploits the discovery that many human proteins are an abundant source of potential CPP sequences which are reliably predicted using QSAR algorithms or other methods. Subsequently, microwave-enhanced solid phase peptides synthesis provides a high-throughput source of novel proteomimetic CPPs for screening purposes. By focussing upon cationic helical domains, often located within the molecular interfaces that facilitate PPIs, bioportides which act by a dominant-negative mechanism at such sites can be reliably identified within small number libraries of CPPs. Protocols that employ fluorescent peptides, routinely prepared by N-terminal acylation with carboxytetramethylrhodamine, further enable both the quantification of cellular uptake kinetics and the identification of specific site(s) of intracellular accretion. Chemical modifications of linear peptides, including strategies to promote and stabilise helicity, are compatible with the synthesis of second-generation bioportides with improved drug-like properties to further exploit the inherent selectivity of biologics

Distinct Behaviour of the Homeodomain Derived Cell Penetrating Peptide Penetratin in Interaction with Different Phospholipids

Penetratin is a protein transduction domain derived from the homeoprotein Antennapedia. Thereby it is currently used as a cell penetrating peptide to introduce diverse molecules into eukaryotic cells, and it could also be involved in the cellular export of transcription factors. Moreover, it has been shown that it is able to act as an antimicrobial agent. The mechanisms involved in all these processes are quite controversial.In this article, we report spectroscopic, calorimetric and biochemical data on the penetratin interaction with three different phospholipids: phosphatidylcholine (PC) and phosphatidylethanolamine (PE) to mimic respectively the outer and the inner leaflets of the eukaryotic plasma membrane and phosphatidylglycerol (PG) to mimic the bacterial membrane. We demonstrate that with PC, penetratin is able to form vesicle aggregates with no major change in membrane fluidity and presents no well defined secondary structure organization. With PE, penetratin aggregates vesicles, increases membrane rigidity and acquires an α-helical structure. With PG membranes, penetratin does not aggregate vesicles but decreases membrane fluidity and acquires a structure with both α-helical and β–sheet contributions.These data from membrane models suggest that the different penetratin actions in eukaryotic cells (membrane translocation during export and import) and on prokaryotes may result from different peptide and lipid structural arrangements. The data suggest that, for eukaryotic cell penetration, penetratin does not acquire classical secondary structure but requires a different conformation compared to that in solution