ABSTRACT
N-Methylpyrrolidone is a solvent molecule which has been shown to compete with acetyl-lysine-containing peptides for binding to bromodomains. From crystallographic studies, it has also been shown to closely mimic the acetamide binding motif in several bromodomains, but has not yet been directly pursued as a fragment in bromodomain inhibition. In this paper, we report the elaboration of N-methylpyrrolidone as a potential lead in fragment-based drug design. Firstly, N-methylpyrrolidone was functionalised to provide points for chemical elaboration. Then, the moiety was incorporated into analogues of the reported bromodomain inhibitor, Olinone. X-ray crystallography revealed that the modified analogues showed comparable binding affinity and structural mimicry to Olinone in the bromodomain binding site.
Subject(s)
Cell Cycle Proteins/chemistry , Drug Design , Pyrrolidinones/chemical synthesis , Transcription Factors/chemistry , Binding Sites , Cell Cycle Proteins/metabolism , Crystallography, X-Ray , Fluorescence Resonance Energy Transfer , Gene Expression Regulation/drug effects , Humans , Models, Molecular , Molecular Structure , Protein Binding , Protein Conformation , Pyrrolidinones/chemistry , Structure-Activity Relationship , Transcription Factors/metabolismABSTRACT
Kisspeptin analogues with improved metabolic stability may represent important ligands in the study of the kisspeptin/KISS1R system and have therapeutic potential. In this paper we assess the activity of known and novel kisspeptin analogues utilising a dual luciferase reporter assay in KISS1R-transfected HEK293T cells. In general terms the results reflect the outcomes of other assay formats and a number of potent agonists were identified among the analogues, including ß(2) -hTyr-modified and fluorescently labelled forms. We also showed, by assaying kisspeptin in the presence of protease inhibitors, that proteolysis of kisspeptin activity within the reporter assay itself may diminish the agonist outputs. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.
Subject(s)
Amino Acids/chemistry , Kisspeptins/agonists , Receptors, G-Protein-Coupled/metabolism , Fluorescent Dyes/chemistry , HEK293 Cells , Humans , Ligands , Receptors, G-Protein-Coupled/chemistry , Receptors, Kisspeptin-1ABSTRACT
Sialic acids (SA) usually linked to galactose (Gal) in an α2,6- or α2,3-configuration are considered the main cell receptors for influenza viruses, in particular for their hemagglutinins (HA). The typing of influenza virus HA receptor selectivity is relevant for understanding the transmissibility of avian and swine viruses to the human population. In this study we developed a simple and inexpensive gel-capture assay (GCA) of the influenza virus HA receptor-binding selectivity. Its principle is the binding of soluble influenza virus to pentasaccharide analogs, representatives of receptors of human and avian influenza viruses, immobilized on a gel resin. The human and avian analogs consisted of a sialyllactose-N-tetraose c (LSTc) [Neu5Ac(α2,6)Gal(ß1-3)GlcNAc(ß1-3)Gal(ß1-4)Glc] and a sialyllactose-N-tetraose a (LSTa) [Neu5Ac(α2,3)Gal(ß1-3)GlcNAc(ß1-3)Gal(ß1-4)Glc], respectively. Following equilibration, the unbound virus is washed away and the bound one is assayed via HA by densitometry as a function of the analog concentration. Using GCA, the receptor selectivity of three influenza viruses of different HA subtype was investigated. The results showed that the egg-adapted A/California/07/2009 (H1N1) virus exhibited an avian α2,3-linked LSTa selectivity, however, it retained the ability to bind to the α2,6-linked LSTc human receptor analog. Influenza B virus B/Florida/4/2006 showed α2,6-linked LSTc selectivity and a poor α2,3-linked LSTa avidity. The H3N2 virus A/Wisconsin/15/2009 displayed almost comparable avidity for both receptor analogs with a marginally greater α2,3-linked LSTa avidity. The described assay protocol provides a simple and rapid method for the characterization of influenza virus HA receptor binding selectivity.
Subject(s)
Host Specificity , Influenza A virus/physiology , Influenza B virus/physiology , Influenza, Human/virology , Polysaccharides/analysis , Receptors, Virus/analysis , Sialic Acids/analysis , Virology/methods , Animals , Chick Embryo , Humans , Influenza A virus/chemistry , Influenza B virus/chemistry , Influenza, Human/metabolism , Polysaccharides/metabolism , Receptors, Virus/metabolism , Sialic Acids/metabolism , Virology/instrumentationABSTRACT
N-Methylpyrrolidone is one of several chemotypes that have been described as a mimetic of acetyl-lysine in the development of bromodomain inhibitors. In this paper, we describe the synthesis of a 4-phenyl substituted analogue - 1-methyl-4-phenylpyrrolidin-2-one - and the use of aryl substitution reactions as a divergent route for derivatives. Ultimately, this has led to structurally complex, chiral compounds with progressively improved affinity as inhibitors of bromodomain-containing protein 4.
Subject(s)
Cell Cycle Proteins/antagonists & inhibitors , Drug Design , Pyrrolidinones/pharmacology , Transcription Factors/antagonists & inhibitors , Cell Cycle Proteins/metabolism , Crystallography, X-Ray , Dose-Response Relationship, Drug , Fluorescence Resonance Energy Transfer , Humans , Models, Molecular , Molecular Structure , Pyrrolidinones/chemistry , Structure-Activity Relationship , Transcription Factors/metabolismABSTRACT
The number of hemoglobins found in individual mature larvae of several Chironomus species is six to nine. The consistency of this number and of the electrophoretic mobilities of these separable forms seems to indicate the synchronous production of polypeptide chains by several genetic loci.
Subject(s)
Diptera , Hemoglobins , Animals , Electrophoresis , Genetics , In Vitro TechniquesABSTRACT
Different populations of Chironomus tentans, possibly representing geographically isolated races, have two differentiated genic mechanisms of sex determination involving either a dominant male-determining factor in the left arm of chromosome 1 or a dominant female-determining factor at the right tip of chromosome 1. In crosses between these populations, the male-determining factor is epistatic to the female-determining factor. No evidence of intersexuality has been found in such crosses.
Subject(s)
Sex Chromosomes , Sex Determination Analysis , Animals , Chromosome Aberrations , Chromosome Mapping , Crosses, Genetic , Diptera , Female , Fertility , Genes, Dominant , Genotype , Larva , Male , Salivary Glands/cytologyABSTRACT
Patients with motor neuron disease with thyroid disorders have been described, although the relationship between the two conditions is unclear. We treated a patient with amyotrophic lateral sclerosis who also had a follicular adenoma of the thyroid gland. Because thyroid gland plasma membranes contain high concentrations of complex gangliosides, such as GD1b, and some patients with motor neuron disease have IgM antibodies to GD1b, we decided to assay serum from this patient for the presence of antiganglioside antibodies. IgM antibodies to GD1b were detectable at serum dilutions of 1:500 and 1:1000 by enzyme-linked immunosorbent assay. While these titers are less than those usually described in patients with plasma cell dyscrasia, they are well in excess of normal values. Antibody to GM1 was also detectable at a lower (1:100) dilution. We do not know the importance of the anti-GD1b antibodies in this patient, but it is possible that antibodies to GD1b are involved in this and other cases of motor neuron disease associated with thyroid disease.
Subject(s)
Adenoma/immunology , Amyotrophic Lateral Sclerosis/immunology , Antibodies/analysis , Gangliosides/immunology , Thyroid Neoplasms/immunology , Adenoma/complications , Amyotrophic Lateral Sclerosis/complications , G(M1) Ganglioside/immunology , Humans , Male , Middle Aged , Thyroid Neoplasms/complicationsABSTRACT
A simple method is described for the application onto HPLC columns of very crude or alternatively poorly soluble polypeptide samples prior to their chromatographic purification. The procedure involves the batch pre-adsorption of the crude polypeptide mixture from a dilute solution onto an appropriate preparative-grade chromatographic adsorbent, removal of the solvent by rotary evaporation or lyophilisation and then dry-packing the pre-adsorbed chromatographic material into guard column cartridges of suitable dimensions. The polypeptide products can then be eluted either by isocratic or gradient elution methods through the cartridge coupled in tandem with prepacked semi-preparative HPLC columns. This method has been successfully utilised for the routine RP-HPLC purification of polar and hydrophobic polypeptides prepared by solid phase peptide synthesis (SPPS) methods as well as peptide derivatives and intermediates used as part of SPPS procedures.
Subject(s)
Peptides/analysis , Adsorption , Amino Acid Sequence , Chromatography, High Pressure Liquid/methods , Molecular Sequence DataABSTRACT
infective larvae of Dipetalonema viteae produced infections in Mongolian jirds (Meriones unguiculatus) after storage of infected ticks (Ornithodoros tartakovskyi) in the presence of dimethyl sulfoxide (DMSO, 5%) for 7 or 595 days in liquid nitrogen (-196 C). Infectivity of these larvae was only partially impaired. Microfilaremias of test jirds were generally lower than those of control jirds given nonfrozen larvae; however, the majority of test jirds developed microfilarial counts suitable for use in infecting ticks. In contradistinction, larvae frozen free of the tick failed to retain infectivity. Apparently the tick, in conjunction with DMSO, protects the larvae during freezing and thawing.
Subject(s)
Dipetalonema/pathogenicity , Preservation, Biological , Animals , Blood/parasitology , Dipetalonema Infections/parasitology , Freezing , Gerbillinae , Larva/pathogenicity , Male , Nitrogen , Ticks/parasitology , Time FactorsABSTRACT
As the numbers of women of childbearing age in the workforce continue to increase, some employers and employees have viewed maternal employment to be incompatible with breast-feeding. This qualitative study investigated factors that hindered as well as facilitated breast-feeding in the workplace. Findings suggest breast-feeding and employment can and should be compatible.
Subject(s)
Adaptation, Psychological , Breast Feeding , Mothers/psychology , Women, Working/psychology , Workplace , Adolescent , Adult , Female , Humans , Infant , Organizational Culture , Social Support , Surveys and QuestionnairesABSTRACT
Asthma, a costly, common chronic condition, needs to be managed proactively rather than reactively. Nurses need to teach children and parents how to recognize and avoid asthma triggers. Asthma management is improved when nurses communicate with the child, parents, and school personnel.
Subject(s)
Asthma/nursing , Asthma/prevention & control , Patient Education as Topic , School Nursing , Adult , Child , Environment , Humans , Parents/educationABSTRACT
Faced with higher medical costs and increased insurance premiums, corporations are focusing on health promotion and wellness. With increasing numbers of women in the workforce, corporations have identified the need for prenatal programs. By developing, initiating, and evaluating outcome-based prenatal programs nurses can target the health care needs of this select population. One such program documented several outcomes including improved employee health and an 86% reduction in maternal/newborn costs.
Subject(s)
Maternal-Child Nursing/organization & administration , Occupational Health Services/organization & administration , Outcome and Process Assessment, Health Care/organization & administration , Prenatal Care/organization & administration , Cost-Benefit Analysis , Female , Health Services Needs and Demand , Health Status , Humans , Pregnancy , Program Development , Program EvaluationABSTRACT
Recent results using animal models of inflammatory skin conditions have shown that blockers of the voltage-gated potassium channel, Kv1.3 hold great promise for clinical utility. Kv1.3 blockers act as immunosuppressants by modulating the various subsets of inflammatory T and B cells involved in autoimmune disorders. While peptidic inhibitors based on naturally occurring venoms demonstrate potent and selective Kv1.3 blockade, these require parenteral administration and may face potential immunogenicity problems. Small molecule blockers show considerable diversity, however selectivity over other Kv1-family channels has been difficult to achieve. More recent advances have added to the evidence that Kv1.3 channels are a suitable therapeutic target and that the development of novel and selective agents will herald new drugs for inflammatory skin disorders.